HK1118287A - Modulators of atp-binding cassette transporters - Google Patents

Description

Modulators of ATP-binding cassette transporters

Cross Reference to Related Applications

The benefit of U.S. provisional application No.60/683,982 entitled "modulators of ATP-binding cassette transporters", filed 2005, 5, 24, § 119, herein under 35u.s.c. The entire contents of each of the aforementioned prior applications are incorporated herein by reference.

Technical Field

The present invention relates to modulators of ATP-binding cassette ('ABC') transporters or fragments thereof, including cystic fibrosis transmembrane conductance regulator ('CFTR'), compositions thereof, and methods therewith. The invention also relates to methods of treating ABC transporter mediated diseases using such modulators.

Background

ABC transporters are a family of membrane transporters that regulate the transport of a variety of pharmacological components, potentially toxic drugs and xenobiotics, as well as anions. ABC transporters are homologous membrane proteins that bind and utilize cellular Adenosine Triphosphate (ATP) for their specific activity. Some of these transporters are found to be multidrug resistance proteins (like MDR1-P glycoprotein or multidrug resistance protein MRP1) that are chemotherapeutics for defense of malignant cancer cells. To date, 48 ABC transporters have been identified, and are classified into 7 families based on their sequence identity and function.

ABC transporters play a variety of important physiological roles in the body and provide defense against harmful environmental compounds. Because of this, they represent important potential drug targets for the treatment of diseases associated with defects in the transporter, prevention of drug transport out of target cells, and intervention in other diseases in which modulation of ABC transporter activity may be beneficial.

One ABC transporter family member commonly associated with disease is the cAMP/ATP-mediated anion channel CFTR. CFTR is expressed in a variety of cell types, including absorptive and secretory epithelial cells, where it regulates the transmembrane flow of anions as well as the activity of other ion channels and proteins. In epithelial cells, normal functioning of CFTR is critical to maintain electrolyte transport throughout the body, including respiratory and digestive tissues. CFTR consists of approximately 1480 amino acids, which encode proteins consisting of tandem repeats of transmembrane domains, each containing six transmembrane helices and a nucleotide binding domain. The two transmembrane domains are linked by a large polarity-regulating (R) -domain, with multiple phosphorylation sites that regulate channel activity and cellular trafficking.

The genes encoding CFTR have been identified and sequenced (see Gregory, R.J. et al (1990) Nature 347: 382. sup. 386; Rich, D.P. et al (1990) Nature 347: 358. sup. 362; Riordan, J.R. et al (1989) Science 245: 1066. sup. 1073). Defects in this gene cause CFTR mutations, leading to cystic fibrosis ("CF"), the most common fatal genetic disease in humans. Cystic fibrosis affects approximately one two thousand five percent of U.S. newborns. Up to ten million people in the entire U.S. population carry a single copy of a defective gene with no apparent disease effects. In contrast, individuals with two copies of the CF-associated gene suffer from the debilitating and fatal effects of CF, including chronic lung disease.

In cystic fibrosis patients, mutations in the respiratory epithelium by endogenously expressed CFTR cause reduced apical anion secretion, resulting in an imbalance in ionic and humoral transport. The resulting anion transport disease contributes to enhanced mucus accumulation in the lung and concomitant microbial infection, ultimately leading to death in CF patients. In addition to respiratory disease, CF patients often suffer from gastrointestinal problems and pancreatic insufficiency, which if left untreated, leads to death. In addition, most cystic fibrosis males are infertile and cystic fibrosis females have reduced fertility. In contrast to the severe effects of two copies of the CF-associated gene, individuals with a single copy of the CF-associated gene exhibit increased resistance to cholera and to dehydration due to diarrhea-perhaps explaining the relatively high frequency of the CF gene in the population.

Sequence analysis of the CFTR gene of the CF chromosome has revealed a number of causative mutations (Cutting, G.R. et al (1990) Nature 346: 366-. More than 1000 pathogenic CF gene mutations have been identified to date (http:// www.genet.sickkids.on.ca/cftr /). The most common mutation is the deletion of phenylalanine at amino acid sequence 508 of CFTR, commonly referred to as Δ F508-CFTR. This mutation occurs in approximately 70% of cystic fibrosis cases and is associated with severe disease.

Deletion of residue 508 in Δ F508-CFTR prevents the nascent protein from folding correctly. This results in the inability of the mutein to exit the ER and be transported to the plasma membrane. As a result, the number of channels in the membrane is much less than in cells expressing wild-type CFTR. In addition to reduced trafficking, mutations also lead to defective channel gating. In summary, a reduced number of channels and defective gating in the membrane leads to reduced anion transport across the epithelium, resulting in defective ion and fluid transport (Quinton, P.M (1990), FASEB J.4: 2709-. However, studies have shown that the reduction in the amount of AF 508-CFTR in the membrane is functional, although less than wild-type CFTR (Dalemans et al (1991), Nature Lond.354: 526-. In addition to af 508-CFTR, other pathogenic CFTR mutations that result in defective trafficking, synthesis, and/or channel gating may be upregulated or downregulated to alter anion secretion and alter disease progression and/or severity.

Although CFTR transports a variety of molecules in addition to anions, it is clear that this effect (transport of anions) represents one of the important mechanisms of transporting ions and water across the epithelium. Other elements include epithelial Na⁺Channel, ENaC, Na⁺/2Cl^-/K⁺Cotransporter, Na⁺-K⁺ATP-ase Pump and basolateral Membrane K⁺Channels, which are responsible for the uptake of chloride into cells.

These elements work together to achieve targeted transport across the epithelium via their selective expression and localization within the cell. By ENaC and CF present on the top membraneTR and Na expressed on the basolateral surface of the cell⁺-K⁺ATP-ase Pump with Cl^-The coordinated activity of the channels, the absorption of chloride takes place. Secondary active transport of chloride from the luminal side causes accumulation of intracellular chloride, which can then passively pass through Cl^-The channel leaves the cell, resulting in vector transport. Na (Na)⁺/2Cl^-/K⁺Cotransporter, Na⁺-K⁺ATP-ase Pump and basolateral Membrane K⁺The arrangement of the channels on the outside surface of the substrate and the CFTR on the luminal side coordinate the secretion of chloride via the CFTR on the luminal side. Because water may never transport itself actively, its flow across the epithelium relies on a slight transepithelial osmotic gradient generated by the bulk flow of sodium and chlorine.

In addition to cystic fibrosis, modulation of CFTR activity may also be beneficial in other diseases not directly caused by CFTR mutations, such as secretory diseases and other protein folding diseases mediated by CFTR. These diseases include, but are not limited to, Chronic Obstructive Pulmonary Disease (COPD), dry eye disease, and sjogren's syndrome.

COPD is characterized by airflow limitation, which is progressive and not fully reversible. Airflow limitation is due to mucus hypersecretion, emphysema, and bronchiolitis. Activators of mutant or wild-type CFTR offer the potential treatment of mucus hypersecretion and reduced mucociliary clearance rates common in COPD. In particular, increasing anion secretion across CFTR may facilitate fluid transport into airway surface fluid to hydrate mucus and optimize fluid viscosity around cilia. This will result in an enhanced mucociliary clearance rate and a reduction in symptoms associated with COPD. Dry eye disease is characterized by decreased tear production and abnormal tear film lipid, protein and mucin behavior. There are many causes of dry eye, some of which include age, Lasik eye surgery, arthritis, medications, chemical/thermal burns, allergies, and diseases such as cystic fibrosis and sjogren's syndrome. Increasing anion secretion via CFTR will enhance transport of body fluids from corneal endothelial cells and secretory glands surrounding the eye to increase hydration of the cornea. This will help to alleviate symptoms associated with dry eye. Sjogren's syndrome is an autoimmune disease in which the immune system attacks glands that produce water everywhere in the body, including the eye, mouth, skin, respiratory tissues, liver, vagina, and intestine. Symptoms include dry eyes, mouth and vagina, and lung disease. The disease is also associated with rheumatoid arthritis, systemic lupus, systemic sclerosis and polymyositis/dermatomyositis. Defective protein trafficking is believed to lead to the disease, and treatment options are limited. Modulators of CFTR activity may hydrate various organs affected by the disease, helping to ameliorate the associated symptoms.

As discussed above, it is believed that the deletion of residue 508 in af 508-CFTR prevents the nascent protein from folding correctly, resulting in the inability of this mutein to exit the ER and be transported to the plasma membrane. As a result, insufficient amounts of mature protein are present at the plasma membrane and chloride transport in epithelial tissues is significantly reduced. In fact, this cellular phenomenon of defective ER processing of ABC transporters by the ER machinery has been shown to underlie not only CF diseases, but a wide range of other isolated and genetic diseases. Two possible modes of failure of the ER machinery are either loss of coupling to the ER export of proteins, causing degradation, or ER accumulation of these defective/misfolded proteins [ Aridor M, et al, Nature med.,5(7)，pp 745-751(1999)；Shastry，B.S.，et al.，Neurochem.International，43，pp 1-7(2003)；Rutishauser，J.，et al.，Swiss Med Wkly，132，pp 211-222(2002)；Morello，JP et al.，TIPS，21，pp.466-469(2000)；Bross P.，et al.，Human Mut.，14，pp.186-198(1999)]. The diseases associated with the former class of ER failure are cystic fibrosis (caused by misfolded AF 508-CFTR, as discussed above), hereditary emphysema (caused by a 1-antitrypsin non-Piz variant), hereditary hemochromatosis, coagulation-fibrinolysis defects (e.g., protein C defect), hereditary angioedema type 1, lipid processing defects (e.g., familial hypercholesterolemia), chylomicronemia type 1, beta-lipoproteinemia, lysosomal storage diseases (e.g., I-cell disease/pseudo-Hurler), mucopolysaccharidosis (processed by lysosomes)Enzyme induced), Sandhof/Tay-Sachs (induced by beta-hexosaminidase), Crigler-Najjar type II (induced by UDP-glucuronyl-sialyc-transferase), polyendocrinopathy/hyperinsulinemia, diabetes (induced by insulin receptor), Laron dwarfism (induced by growth hormone receptor), myeloperoxidase deficiency, primary hypoparathyroidism (induced by prepro-parathyroid hormone), melanoma (induced by tyrosinase). Diseases associated with the latter ER failure are glycan disease type CDG 1, hereditary emphysema (caused by alpha 1-antitrypsin PiZ variants), congenital hyperthyroidism, osteogenesis imperfecta (caused by procollagen types I, II, IV), hereditary hypofibrinogenemia (caused by fibrinogen), ACT deficiency (caused by alpha 1-anti-lactoproteinase), Diabetes Insipidus (DI), neurophysin-carrying DI (caused by vasopressin/V2-receptor), nephrogenic DI (caused by aquaporin II), Charpy-Marek's syndrome (caused by peripheral myelin protein 22), Palmer's disease, neurodegenerative diseases (e.g. Alzheimer's disease (caused by beta and presenilin), Parkinson's disease, amyotrophic lateral sclerosis, progressive supranuclear palsy, Pick's disease), several polyglutamine neurological disorders (e.g., huntington's disease, spinocerebellar ataxia type I, spinal and bulbar muscular atrophy, dentatorubral pallidoluysian, and myotonic dystrophy) and spongiform encephalopathies (e.g., hereditary creutzfeldt-jakob disease (caused by prion protein processing defects), fabry disease (caused by lysosomal α -galactosidase a), and stewartz's syndrome (caused by Prp processing defects)).

In addition to upregulation of CFTR activity, reduction of CFTR secretory anions may also be beneficial in the treatment of secretory diarrhoea, where epithelial water transport is dramatically increased as a result of secretagogue activated chloride transport. This mechanism involves elevation of cAMP and stimulation of CFTR.

Although there are a number of causes of diarrhea, the major consequences of diarrheal disease resulting from excessive chloride transport are common, including dehydration, acidosis, reduced growth and death.

Acute and chronic diarrhea represent a major medical problem in many parts of the world. Diarrhea is both a significant cause of malnutrition and a leading cause of death (5,000,000 deaths per year) in children less than five years of age.

Secretory diarrhoea is also a dangerous condition in patients with acquired immunodeficiency syndrome (AIDS) and chronic Inflammatory Bowel Disease (IBD). Each year, sixteen million people traveling from industrialized countries to developing countries suffer from diarrhea, the severity and number of cases of diarrhea varying depending on the country and region of travel.

Diarrhea in livestock and pets, such as cattle, pigs, horses, sheep, goats, cats and dogs, also known as scours, is a major cause of death in these animals. Diarrhea can be caused by any major transition, such as weaning or physical movement, and in response to a variety of bacterial or viral infections, typically occurring within the first few hours of the animal's life.

The most common diarrheagenic bacterium is enterotoxigenic Escherichia coli (ETEC), with K99 cilium antigen. Common viral causes of diarrhea include rotavirus and coronavirus. Other infectious agents include cryptosporidium, giardia lamblia, and salmonella, among others.

Symptoms of rotavirus infection include excretion of watery feces, dehydration, and weakness. Coronavirus causes more serious neonatal disease with a higher mortality rate than rotavirus infection. Often, however, young animals may be infected with more than one virus or a combination of viruses and bacterial microorganisms at the same time. This dramatically increases the severity of the disease.

Thus, there is a need for modulators of ABC transporter activity and compositions thereof that can be used to modulate the activity of ABC transporters in the cell membrane of a mammal.

There is a need for methods of treating ABC transporter mediated diseases using modulators of ABC transporter activity.

There is a need for methods of modulating ABC transporter activity in mammalian cell membranes ex vivo.

There is a need for modulators of CFTR activity that can be used to modulate the activity of CFTR in mammalian cell membranes.

There is a need for methods of treating CFTR mediated diseases using such modulators of CFTR activity.

There is a need for methods of modulating CFTR activity in mammalian cell membranes ex vivo.

Disclosure of Invention

It has now been found that the compounds of the present invention, and pharmaceutically acceptable salts thereof, are useful as modulators of ABC transporter activity, e.g., CFTR activity. These compounds have the general formula I:

wherein ring a and substituents Ra, Rb, Rc and Rd are as follows.

These compounds and pharmaceutically acceptable compositions are useful for treating or lessening the severity of a variety of diseases, disorders or conditions, including, but not limited to, cystic fibrosis, hereditary emphysema, hereditary hemochromatosis, coagulation-fibrinolysis deficiency (e.g., protein C deficiency), hereditary angioedema type 1, lipid processing deficiency (e.g., familial hypercholesterolemia), chylomicronemia type 1, beta-lipoproteinemia, lysosomal storage diseases (e.g., I-cell disease/pseudo Hurler), mucopolysaccharidoses, Sandhof/Tay-Sachs, Crigler-Najjar type II, polyendocrinopathy/hyperinsulinemia, diabetes, Laron dwarfism, myeloperoxidase deficiency, primary hypoparathyroidism, melanoma, polyoses CDG type 1, hereditary emphysema, congenital hyperthyroidism, Osteogenesis imperfecta, hereditary hypofibrinogenemia, ACT deficiency, Diabetes Insipidus (DI), neurophysin-carrying DI, nephrogenic DI, Charcot-Marie-tooth syndrome, Palmer's disease, neurodegenerative diseases (e.g., Alzheimer's disease, Parkinson's disease, amyotrophic lateral sclerosis, progressive supranuclear palsy, pick's disease), several polyglutamine neurological disorders (e.g., Huntington's disease, spinocerebellar ataxia type I, spinal and bulbar muscular atrophy, dentate red nucleus pallidoluysian, and myotonic dystrophy), and spongiform encephalopathies (e.g., hereditary Creutzfeldt-Jakob disease, Fabry's disease, and Stess-Sjogren syndrome), COPD, dry eye disease, and Sjogren's disease.

Detailed description of the invention

I. Definition of

The following definitions apply herein, unless otherwise indicated.

The term "ABC-transporter" as used herein means an ABC-transporter protein or a fragment thereof comprising at least one binding domain, wherein said protein or fragment thereof is present in vivo or in vitro. The term "binding domain" as used herein denotes a domain on an ABC-transporter that is capable of binding to a modulator. See, e.g., Hwang, t.c.et al, j.gen.physiol. (1998): 111(3),477-90.

The term "CFTR" as used herein means a cystic fibrosis transmembrane regulator or partial or complete mutant thereof having regulator activity, including but not limited to Δ F508-CFTR and G551D-CFTR (see, e.g., http:// www.genet.sickkids.on.ca/CFTR/for CFTR mutations).

The term "modulate" as used herein means to increase or decrease, e.g., activity, to a measurable amount. Compounds that modulate ABC transporter activity, e.g., CFTR activity, by increasing the activity of ABC transporters, e.g., CFTR anion channels, are referred to as agonists. Compounds that modulate ABC transporter activity, e.g., CFTR activity, by decreasing the activity of ABC transporters, e.g., CFTR anion channels, are referred to as antagonists. Agonists act on ABC transporters, such as CFTR anion channels, increasing the ability of the receptor to transduce an intracellular signal in response to endogenous ligand binding. Antagonists act on ABC transporters, such as CFTR, and compete with endogenous ligands or substrates for binding sites on the receptor, reducing the ability of the receptor to transduce intracellular signals in response to endogenous ligand binding.

The phrase "treating or reducing the severity of an ABC transporter mediated disease" refers to both treating a disease caused directly by ABC transporter and/or CFTR activity and alleviating the symptoms of a disease not caused directly by ABC transporter and/or CFTR anion channel activity. Examples of diseases whose symptoms may be affected by ABC transporter and/or CFTR activity include, but are not limited to, cystic fibrosis, hereditary emphysema, hereditary hemochromatosis, coagulation-fibrinolysis deficiencies (e.g., protein C deficiency), hereditary angioedema type 1, lipid processing deficiencies (e.g., familial hypercholesterolemia), chylomicronemia type 1, β -lipoproteinemia, lysosomal storage diseases (e.g., I-cell disease/pseudo Hurler), mucopolysaccharidoses, Sandhof/Tay-Sachs, Crigler-Najjar type II, polyendocrinopathy/hyperinsulinemia, diabetes, Laron dwarfism, myeloperoxidase deficiency, primary hypoparathyroidism, melanoma, polyoses type CDG 1, hereditary emphysema, congenital hyperthyroidism, osteogenesis imperfecta, hereditary hypoproteinemia, ACT deficiency, Diabetes Insipidus (DI), posterior leaflet hormone-transporter DI, nephrogenic DI, Charcot-Marie-Tourette syndrome, Palmer's disease, neurodegenerative diseases (e.g., Alzheimer's disease, Parkinson's disease, amyotrophic lateral sclerosis, progressive supranuclear palsy, pick's disease), several polyglutamine neurological disorders (e.g., Huntington's disease, spinocerebellar ataxia type I, spinal and bulbar muscular atrophy, dentate red nucleus pallidoluysian, and myotonic dystrophy), and spongiform encephalopathies (e.g., hereditary Creutzfeldt-Jakob disease, Fabry's disease, and Sjogren's syndrome), COPD, dry eye disease, and Sjogren's disease.

For the purposes of the present invention, the chemical elements conform to the Periodic Table of the elements, CAS version, Handbook of Chemistry and Physics, 75th ed. In addition, the general principles of Organic Chemistry are described in "Organic Chemistry", ThomasSorrell, University Science Books, Sausolato: 1999, and "March's Advanced Organic Chemistry", 5th Ed., Ed.: smith, m.b. and March, j., John Wiley & Sons, New York: 2001, the entire contents of which are incorporated herein by reference.

The term "aliphatic" as used herein encompasses the terms alkyl, alkenyl, alkynyl, each of which is optionally substituted as described below.

As used herein, "alkyl" refers to a saturated aliphatic hydrocarbon group containing 1-8 (e.g., 1-6 or 1-4) carbon atoms. The alkyl group may be linear or branched. Examples of alkyl groups include, but are not limited to, methyl, ethyl, propyl, isopropyl, butyl, isobutyl, sec-butyl, tert-butyl, n-pentyl, n-heptyl, or 2-ethylhexyl. Alkyl groups may be (that is to say are optionally) substituted by one or more substituents, for example halogen, cycloaliphatic (for example cycloalkyl or cycloalkenyl), heterocycloaliphatic (for example heterocycloalkyl or heterocycloalkenyl), aryl, heteroaryl, alkoxy, aroyl, heteroaroyl, acyl (for example (aliphatic) carbonyl, (cycloaliphatic) carbonyl or (heterocycloaliphatic) carbonyl), nitro, cyano, acylamino (for example (cycloalkylalkyl) carbonylamino, arylcarbonylamino, aralkylcarbonylamino, (heterocycloalkyl) carbonylamino, (heterocycloalkylalkyl) carbonylamino, heteroarylcarbonylamino, heteroaralkylcarbonylamino, alkylaminocarbonyl, cycloalkylaminocarbonyl, heterocycloalkylaminocarbonyl, arylaminocarbonyl or heteroarylaminocarbonyl), amino (for example aliphatic amino, cycloaliphatic amino or heterocycloaliphatic amino), Sulfonyl (e.g. aliphatic-SO)₂-), sulfinyl, sulfanyl, sulfoxy (sulfoxy), urea, thiourea, sulfamoyl, sulfonylamino, oxo, carboxyl, carbamoyl, cycloaliphatic oxy, heterocycloaliphatic oxy, aryloxy, heteroaryloxy, aralkyloxy, heteroarylalkoxy, alkoxycarbonyl, alkylcarbonyloxy or hydroxyl. Some examples of substituted alkyl groups include, without limitation, carboxyalkyl (e.g., HOOC-alkyl, alkoxycarbonylalkyl, and alkylcarbonyloxyalkyl), cyanoalkyl, hydroxyalkaneAlkyl, alkoxyalkyl, acylalkyl, aralkyl, (alkoxyaryl) alkyl, (sulfonylamino) alkyl (e.g., (alkyl-SO)₂-amino) alkyl), aminoalkyl, amidoalkyl, (cycloaliphatic) alkyl or haloalkyl.

"alkenyl" as used herein, means an aliphatic carbon group containing 2 to 8 (e.g., 2 to 6 or 2 to 4) carbon atoms and at least one double bond. Like the alkyl group, the alkenyl group may be linear or branched. Examples of alkenyl groups include, but are not limited to, allyl, isopropenyl, 2-butenyl, and 2-hexenyl. The alkenyl group may be optionally substituted with one or more substituents, such as halogen, cycloaliphatic (e.g., cycloalkyl or cycloalkenyl), heterocycloaliphatic (e.g., heterocycloalkyl or heterocycloalkenyl), aryl, heteroaryl, alkoxy, aroyl, heteroaroyl, acyl (e.g., (aliphatic) carbonyl, (cycloaliphatic) carbonyl, or (heterocycloaliphatic) carbonyl), nitro, cyano, amido (e.g., (cycloalkylalkyl) carbonylamino, arylcarbonylamino, aralkylcarbonylamino, (heterocycloalkyl) carbonylamino, heteroarylcarbonylamino, heteroaralkylcarbonylamino, alkylaminocarbonyl, cycloalkylaminocarbonyl, heterocycloalkylaminocarbonyl, arylaminocarbonyl or heteroarylaminocarbonyl), amino (e.g., aliphatic amino, cycloaliphatic amino, heterocycloaliphatic amino, or aliphatic sulfonylamino), Sulfonyl (e.g. alkyl-SO)₂-, cycloaliphatic radical-SO₂-or aryl-SO₂-), sulfinyl, sulfanyl, sulfoxy, urea, thiourea, sulfamoyl, sulfonylamino, oxo, carboxy, carbamoyl, cycloaliphatic oxy, heterocycloaliphatic oxy, aryloxy, heteroaryloxy, aralkoxy, heteroarylalkoxy, alkoxycarbonyl, alkylcarbonyloxy or hydroxy. Some examples of substituted alkenyl groups include, without limitation, cyanoalkenyl, alkoxyalkenyl, acylalkenyl, hydroxyalkenyl, aralkenyl, (alkoxyaryl) alkenyl, (sulfonylamino) alkenyl (e.g., (alkyl-SO)₂-amino) alkenyl), aminoalkenyl, amidoalkenyl, (cycloaliphatic) alkenyl, or haloalkenyl.

As used herein, "alkynyl" means containing 2-8 (e.g., 2-6) substituentsOr 2-4) carbon atoms and at least one triple bond. The alkynyl group may be linear or branched. Examples of alkynyl groups include, but are not limited to, propargyl and butynyl. Alkynyl groups may be optionally substituted with one or more substituents, such as aroyl, heteroaroyl, alkoxy, cycloalkoxy, heterocycloalkoxy, aryloxy, heteroaryloxy, aralkoxy, nitro, carboxy, cyano, halogen, hydroxy, sulfo, mercapto, sulfanyl (e.g., aliphatic sulfanyl or cycloaliphatic sulfanyl), sulfinyl (e.g., aliphatic sulfinyl or cycloaliphatic sulfinyl), sulfonyl (e.g., aliphatic-SO)₂-, aliphatic amino-SO₂Or cycloaliphatic radicals-SO₂-), acylamino (e.g. aminocarbonyl, alkylaminocarbonyl, alkylcarbonylamino, cycloalkylaminocarbonyl, heterocycloalkylaminocarbonyl, cycloalkylcarbonylamino, arylaminocarbonyl, arylcarbonylamino, aralkylcarbonylamino, (heterocycloalkyl) carbonylamino, (cycloalkylalkyl) carbonylamino, heteroaralkylcarbonylamino, heteroarylcarbonylamino or heteroarylaminocarbonyl), urea, thiourea, sulfamoyl, sulfonylamino, alkoxycarbonyl, alkylcarbonyloxy, cycloaliphatic, heterocycloaliphatic, aryl, heteroaryl, acyl (e.g. (cycloaliphatic) carbonyl or (heterocycloaliphatic) carbonyl), amino (e.g. aliphatic amino), sulfoxy, oxo, carboxyl, carbamoyl, (cycloaliphatic) oxy, (heterocycloaliphatic) oxy or (heteroaryl) alkoxy.

"amido" as used herein encompasses "aminocarbonyl" and "carbonylamino". These terms, when used alone or in combination with another group, denote an amido group, e.g., -N (R) when used terminally^X)-C(O)-R^Yor-C (O) -N (R)^X)₂And when used internally-C (O) -N (R)^X) -or-N (R)^X) -C (O) -, wherein R^XAnd R^YIs defined as follows. Examples of acylamino groups include alkylamido (e.g., alkylcarbonylamino or alkylaminocarbonyl), (heterocycloaliphatic) acylamino, (heteroaralkyl) acylamino, (heteroaryl) acylamino, (heterocycloalkyl) alkylamido, arylamido, aralkylamido, (cycloalkyl) alkylamido or cycloalkylacylAn amino group.

As used herein, "amino" refers to-NR^XR^YWherein each R is^XAnd R^YIndependently hydrogen, alkyl, cycloaliphatic, (cycloaliphatic) aliphatic, aryl, araliphatic, heterocycloaliphatic, (heterocycloaliphatic) aliphatic, heteroaryl, carboxy, sulfanyl, sulfinyl, sulfonyl, (aliphatic) carbonyl, (cycloaliphatic) carbonyl, ((cycloaliphatic) aliphatic) carbonyl, arylcarbonyl, (araliphatic) carbonyl, (heterocycloaliphatic) carbonyl, ((heterocycloaliphatic) aliphatic) carbonyl, (heteroaryl) carbonyl, or (heteroarylaliphatic) carbonyl, each as defined herein and optionally substituted. Examples of the amino group include an alkylamino group, a dialkylamino group, or an arylamino group. When the term "amino" is not a terminal group (e.g., alkylcarbonylamino), it is represented by-NR^X-is represented by. R^XHave the same meaning as defined above.

As used herein, "aryl" used alone or as part of a larger group "aralkyl", "aralkoxy", or "aryloxyalkyl" refers to monocyclic (e.g., phenyl), bicyclic (e.g., indenyl, naphthyl, tetrahydronaphthyl, tetrahydroindenyl), and tricyclic (e.g., fluorenyl, tetrahydrofluorenyl, tetrahydroanthracenyl, anthracenyl) ring systems in which the monocyclic ring system is aromatic or at least one of the bicyclic or tricyclic ring systems is aromatic. Bicyclic and tricyclic groups include benzo-fused 2-3 membered carbocyclic rings. For example, the benzo-fused group includes a group with two or more C_4-8Carbocyclic groups fused phenyl. Aryl groups are optionally substituted with one or more substituents, including aliphatic groups (e.g., alkyl, alkenyl, or alkynyl groups); a cycloaliphatic group; (cycloaliphatic) aliphatic groups; a heterocycloaliphatic group; (heterocycloaliphatic) aliphatic; an aryl group; a heteroaryl group; an alkoxy group; (cycloaliphatic) oxy; (heterocycloaliphatic) oxy; an aryloxy group; a heteroaryloxy group; (araliphatic) oxy; (heteroarylaliphatic) oxy; aroyl; a heteroaroyl group; an amino group; oxo (on a non-aromatic carbocyclic ring of a benzo-fused bicyclic or tricyclic aryl); a nitro group; a carboxyl group; an amido group; acyl (e.g. aliphatic; (cycloaliphatic); (cyclo;) carbonyl; (cyclo)Aliphatic group) carbonyl group; (araliphatic) carbonyl; (heterocycloaliphatic) carbonyl; a ((heterocycloaliphatic) aliphatic) carbonyl group; or a (heteroaraliphatic) carbonyl group); sulfonyl (e.g. aliphatic-SO)₂-or amino-SO₂-) according to the formula (I); sulfinyl (e.g., aliphatic-S (O) -or cycloaliphatic-S (O)); sulfanyl (e.g., aliphatic-S-); a cyano group; halogen; a hydroxyl group; a mercapto group; sulfoxy group; urea; thiourea; a sulfamoyl group; a sulfonylamino group; or a carbamoyl group. Alternatively, the aryl group may be unsubstituted.

Non-limiting examples of substituted aryl groups include haloaryl groups (e.g., mono-, di (e.g., p, m-dihaloaryl) and (trihalo) aryl); (carboxy) aryl groups (e.g., (alkoxycarbonyl) aryl groups, ((aralkyl) carbonyloxy) aryl groups, and (alkoxycarbonyl) aryl groups); (amido) aryl groups (e.g., (aminocarbonyl) aryl, ((alkylamino) alkyl) aminocarbonyl) aryl, (alkylcarbonyl) aminoaryl, (arylaminocarbonyl) aryl, and (((heteroaryl) amino) carbonyl) aryl); aminoaryl (e.g., ((alkylsulfonyl) amino) aryl or ((dialkyl) amino) aryl); (cyanoalkyl) aryl; (alkoxy) aryl; (sulfamoyl) aryl (e.g., (sulfamoyl) aryl); (alkylsulfonyl) aryl; (cyano) aryl; (hydroxyalkyl) aryl; ((alkoxy) alkyl) aryl; (hydroxy) aryl, ((carboxy) alkyl) aryl; ((dialkyl) amino) alkyl) aryl; (nitroalkyl) aryl; ((alkylsulfonyl) amino) alkyl) aryl; ((heterocycloaliphatic) carbonyl) aryl; ((alkylsulfonyl) alkyl) aryl; (cyanoalkyl) aryl; (hydroxyalkyl) aryl; (alkylcarbonyl) aryl; an alkylaryl group; (trihaloalkyl) aryl; p-amino-m-alkoxycarbonylaryl; p-amino-m-cyanoaryl; p-halo-m-aminoaryl; or (m- (heterocycloaliphatic) -o- (alkyl)) aryl.

As used herein, "araliphatic radical" such as "aralkyl" refers to an aliphatic radical (e.g., C) substituted with an aryl radical_1-4Alkyl groups). "aliphatic", "alkyl" and "aryl" are as defined herein. An example of an araliphatic group, such as an aralkyl group, is benzyl.

As used hereinThe "aralkyl group" of (A) represents an alkyl group substituted with an aryl group (e.g., G)_1-4Alkyl groups). Both "alkyl" and "aryl" have been defined above. An example of an aralkyl group is benzyl. Aralkyl groups may be optionally substituted with one or more substituents, such as aliphatic (e.g., alkyl, alkenyl, or alkynyl groups including carboxyalkyl, hydroxyalkyl, or haloalkyl groups such as trifluoromethyl), cycloaliphatic (e.g., cycloalkyl or cycloalkenyl), (cycloalkyl) alkyl, heterocycloalkyl, (heterocycloalkyl) alkyl, aryl, heteroaryl, alkoxy, cycloalkoxy, heterocycloalkoxy, aryloxy, heteroaryloxy, aralkoxy, heteroarylalkoxy, aroyl, heteroaroyl, nitro, carboxyl, alkoxycarbonyl, alkylcarbonyloxy, amido (e.g., aminocarbonyl, alkylcarbonylamino, cycloalkylcarbonylamino, (cycloalkylalkyl) carbonylamino, arylcarbonylamino, aralkylcarbonylamino, (heterocycloalkyl) carbonylamino, heteroarylcarbonylamino, or heteroaralkylcarbonylamino), Cyano, halogen, hydroxy, acyl, mercapto, alkylsulfanyl, sulfoxy, urea, thiourea, sulfamoyl, sulfonylamino, oxo, or carbamoyl.

As used herein, a "bicyclic ring system" includes an 8-12 (e.g., 9, 10, or 11) membered structure that forms two rings, wherein the two rings have at least one atom in common (e.g., 2 atoms in common). Bicyclic ring systems include bicycloaliphatic (e.g., bicycloalkyl or bicycloalkenyl), bicycloheteroaliphatic, bicycloaryl, and bicycloheteroaryl groups.

As used herein, "cycloaliphatic radical" encompasses "cycloalkyl" and "cycloalkenyl", each of which may be optionally substituted as described below.

"cycloalkyl" as used herein means a saturated carbocyclic mono-or bicyclic (fused or bridged) ring of 3 to 10 (e.g., 5 to 10) carbon atoms. Examples of cycloalkyl include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, adamantyl, norbornyl, cubyl, octahydroindenyl, decahydronaphthyl, bicyclo [3.2.1]Octyl, bicyclo [2.2.2]Octyl, bicyclo [3.3.1]Nonyl, bicyclo [3.3.2.]Decyl, bicyclo [2.2.2]Octyl diamondAlkyl, azacycloalkyl or ((aminocarbonyl) cycloalkyl. "cycloalkenyl" as used herein, means a non-aromatic carbocyclic ring of 3 to 10 (e.g., 4 to 8) carbon atoms having one or more double bonds. Examples of cycloalkenyl include cyclopentenyl, 1, 4-cyclohexadienyl, cycloheptenyl, cyclooctenyl, hexahydroindenyl, octahydronaphthyl, cyclohexenyl, cyclopentenyl, bicyclo [2.2.2]Octenyl or bicyclo [3.3.1]Nonenyl. The cycloalkyl or cycloalkenyl groups can be optionally substituted with one or more substituents, such as aliphatic (e.g., alkyl, alkenyl, or alkynyl), cycloaliphatic, (cycloaliphatic) aliphatic, heterocycloaliphatic, (heterocycloaliphatic) aliphatic, aryl, heteroaryl, alkoxy, (cycloaliphatic) oxy, (heterocycloaliphatic) oxy, aryloxy, heteroaryloxy, (araliphatic) oxy, (heteroarylaliphatic) oxy, aroyl, heteroaroyl, amino, amido (e.g., (aliphatic) carbonylamino, (cycloaliphatic) carbonylamino, ((cycloaliphatic) aliphatic) carbonylamino, (aryl) carbonylamino, (araliphatic) carbonylamino, (heterocycloaliphatic) carbonylamino, ((heterocycloaliphatic) aliphatic) carbonylamino, (heteroaryl) carbonylamino, or (heteroarylaliphatic) carbonylamino), Nitro, carboxyl (e.g. HOOC-, alkoxycarbonyl or alkylcarbonyloxy), acyl (e.g. (cycloaliphatic) carbonyl, ((cycloaliphatic) aliphatic) carbonyl, (araliphatic) carbonyl, (heterocycloaliphatic) carbonyl, ((heterocycloaliphatic) aliphatic) carbonyl, or (heteroarylaliphatic) carbonyl), cyano, halogen, hydroxy, mercapto, sulfonyl (e.g. alkyl-SO)₂And aryl-SO₂-), sulfinyl (e.g., alkyl-S (O) -, sulfanyl (e.g., alkyl-S-), sulfoxy, urea, thiourea, sulfamoyl, sulfonylamino, oxo, or carbamoyl.

As used herein, "cyclic group" includes cycloaliphatic, heterocycloaliphatic, aryl, or heteroaryl groups, each of which has been defined above.

The term "heterocycloaliphatic" as used herein encompasses heterocycloalkyl and heterocycloalkenyl, each of which may be optionally substituted as described below.

As used herein, "heterocycle"Alkyl "means a 3-10 membered mono-or bicyclic (fused or bridged) (e.g., 5-to 10-membered mono-or bicyclic) saturated ring structure in which one or more ring atoms is a heteroatom (e.g., N, O, S or a combination thereof). Examples of heterocycloalkyl include piperidyl, piperazinyl, tetrahydropyranyl, tetrahydrofuranyl, 1, 4-dioxolanyl, 1, 4-dithianyl, 1, 3-dioxolanyl, oxazolidinyl, isoxazolidinyl, morpholinyl, thiomorpholinyl, octahydrobenzofuranyl, octahydrobenzopyranyl, octahydrobenzothiopyranyl, octahydroindolyl, octahydropyridinyl, decahydroquinolinyl, octahydrobenzo [ b ] b]Thienyl, 2-oxa-bicyclo [2.2.2]Octyl, 1-aza-bicyclo [2.2.2]Octyl, 3-aza-bicyclo [3.2.1]Octyl and 2, 6-dioxa-tricyclo [3.3.1.0^3，7]Nonyl radical. Monocyclic heterocycloalkyl groups may be fused to phenyl groups, such as tetrahydroisoquinoline. As used herein, "heterocycloalkenyl" refers to a mono-or bicyclic (e.g., 5-to 10-membered mono-or bicyclic) non-aromatic ring structure having one or more double bonds in which one or more ring atoms is a heteroatom (e.g., N, O or S). Monocyclic and bicyclic heteroaliphatic groups are numbered according to standard chemical nomenclature.

The heterocycloalkyl or heterocycloalkenyl group can be optionally substituted with one or more substituents, such as aliphatic (e.g., alkyl, alkenyl, or alkynyl), cycloaliphatic, (cycloaliphatic) aliphatic, heterocycloaliphatic, (heterocycloaliphatic) aliphatic, aryl, heteroaryl, alkoxy, (cycloaliphatic) oxy, (heterocycloaliphatic) oxy, aryloxy, heteroaryloxy, (araliphatic) oxy, (heteroarylaliphatic) oxy, aroyl, heteroaroyl, amino, amido (e.g., (aliphatic) carbonylamino, (cycloaliphatic) carbonylamino, ((cycloaliphatic) aliphatic) carbonylamino, (aryl) carbonylamino, (araliphatic) carbonylamino, (heterocycloaliphatic) carbonylamino, ((heterocycloaliphatic) aliphatic) carbonylamino, (heteroaryl) carbonylamino, or (heteroarylaliphatic) carbonylamino), Nitro, carboxyl (e.g. HOOC-, alkoxycarbonyl or alkylcarbonyloxy), acyl (e.g. (cycloaliphatic) carbonyl, ((cycloaliphatic) aliphatic) carbonyl, (araliphatic) carbonyl, (heterocycloaliphatic) carbonyl, ((heterocycloaliphatic) aliphatic) carbonyl, or (heteroarylaliphatic) carbonyl), nitro, cyano, halogen, hydroxy, mercapto, sulfonyl (e.g. alkylsulfonyl or arylsulfonyl), sulfinyl (e.g. alkylsulfinyl), sulfanyl (e.g. alkylsulfanyl), sulfoxyloxy, urea, thiourea, sulfamoyl, sulfonylamino, oxo, or carbamoyl.

As used herein, "heteroaryl" refers to a monocyclic, bicyclic, or tricyclic ring system having 4 to 15 ring atoms in which one or more ring atoms is a heteroatom (e.g., N, O, S or a combination thereof), wherein the monocyclic ring system is aromatic or at least one ring in the bicyclic or tricyclic ring system is aromatic. Heteroaryl includes benzo-fused ring systems having 2 to 3 rings. For example, benzo-fused groups include benzo-fused to one or two 4-to 8-membered heterocycloaliphatic groups (e.g., indolizinyl, indolyl, isoindolyl, 3H-indolyl, indolinyl, benzo [ b ] furanyl, benzo [ b ] thienyl, quinolinyl, or isoquinolinyl). Examples of some heteroaryl groups are azetidinyl, pyridyl, 1H-indazolyl, furyl, pyrrolyl, thienyl, thiazolyl, oxazolyl, imidazolyl, tetrazolyl, benzofuryl, isoquinolyl, benzothiazolyl, xanthene, thioxanthene, phenothiazine, indoline, benzo [1, 3] dioxole, benzo [ b ] furyl, benzo [ b ] thienyl, indazolyl, benzimidazolyl, benzothiazolyl, purinyl, cinnolinyl, quinolyl, quinazolinyl, cinnolinyl, phthalazinyl, quinazolinyl, quinoxalinyl, isoquinolyl, 4H-quinolizinyl, benzo-1, 2, 5-thiadiazolyl or 1, 8-naphthyridinyl.

Monocyclic heteroaryl groups include, but are not limited to, furyl, thienyl, 2H-pyrrolyl, oxazolyl, thiazolyl, imidazolyl, pyrazolyl, isoxazolyl, isothiazolyl, 1, 3, 4-thiadiazolyl, 2H-pyranyl, 4-H-pyranyl, pyridyl, pyridazinyl, pyrimidinyl, pyrazolyl, pyrazinyl or 1, 3, 5-triazinyl. Monocyclic heteroaryl groups are numbered according to standard chemical nomenclature.

Bicyclic heteroaryl groups include, but are not limited to, indolizinyl, indolyl, isoindolyl, 3H-indolyl, indolinyl, benzo [ b ] furanyl, benzo [ b ] thienyl, quinolinyl, isoquinolinyl, indolizinyl, isoindolyl, indolyl, benzo [ b ] furanyl, benzo [ b ] thienyl, indazolyl, benzimidazolyl, benzothiazolyl, purinyl, 4H-quinolizinyl, quinolinyl, isoquinolinyl, cinnolinyl, phthalazinyl, quinazolinyl, quinoxalinyl, 1, 8-naphthyridinyl, or pteridinyl. Bicyclic heteroaryls are numbered according to standard chemical nomenclature.

Heteroaryl is optionally substituted with one or more substituents, such as aliphatic (e.g., alkyl, alkenyl, or alkynyl); a cycloaliphatic group; (cycloaliphatic) aliphatic groups; a heterocycloaliphatic group; (heterocycloaliphatic) aliphatic; an aryl group; a heteroaryl group; an alkoxy group; (cycloaliphatic) oxy; (heterocycloaliphatic) oxy; an aryloxy group; a heteroaryloxy group; (araliphatic) oxy; (heteroarylaliphatic) oxy; aroyl; a heteroaroyl group; an amino group; oxo (on a non-aromatic carbocyclic or heterocyclic ring of a bicyclic or tricyclic heteroaryl); a carboxyl group; an amido group; acyl (e.g., an aliphatic carbonyl group, (cycloaliphatic) aliphatic carbonyl group, (araliphatic) carbonyl group, (heterocycloaliphatic) aliphatic) carbonyl group, or (heteroarylaliphatic) carbonyl group); sulfonyl (e.g., aliphatic sulfonyl or aminosulfonyl); sulfinyl (e.g., aliphaticsulfinyl); sulfanyl (e.g., aliphatic sulfanyl); a nitro group; a cyano group; halogen; a hydroxyl group; a mercapto group; sulfoxy group; urea; thiourea; a sulfamoyl group; a sulfonylamino group; or a carbamoyl group. Alternatively, the heteroaryl group can be unsubstituted.

Non-limiting examples of substituted heteroaryl groups include (halo) heteroaryl groups (e.g., mono-and di- (halo) heteroaryl); (carboxy) heteroaryl (e.g., (alkoxycarbonyl) heteroaryl); a cyanoheteroaryl group; aminoheteroaryl (e.g., ((alkylsulfonyl) amino) heteroaryl and ((dialkyl) amino) heteroaryl); (amido) heteroaryl groups (e.g., aminocarbonylheteroaryl groups, ((alkylcarbonyl) amino) heteroaryl groups, (((alkyl) amino) alkyl) aminocarbonyl) heteroaryl groups, (((heteroaryl) amino) carbonyl) heteroaryl groups, ((heterocycloaliphatic) carbonyl) heteroaryl groups, and ((alkylcarbonyl) amino) heteroaryl groups); (cyanoalkyl) heteroaryl; (alkoxy) heteroaryl; (sulfamoyl) heteroaryl (e.g., (aminosulfonyl) heteroaryl); (sulfonyl) heteroaryl (e.g., (alkylsulfonyl) heteroaryl); (hydroxyalkyl) heteroaryl; (alkoxyalkyl) heteroaryl; (hydroxy) heteroaryl; ((carboxy) alkyl) heteroaryl; ((dialkyl) amino) alkyl ] heteroaryl, (heterocycloaliphatic) heteroaryl, (cycloaliphatic) heteroaryl, (nitroalkyl) heteroaryl, (((alkylsulfonyl) amino) alkyl) heteroaryl, (alkylsulfonyl) alkyl) heteroaryl, (cyanoalkyl) heteroaryl, (acyl) heteroaryl (e.g., (alkylcarbonyl) heteroaryl), (alkyl) heteroaryl, and (haloalkyl) heteroaryl (e.g., trihaloalkylheteroaryl).

As used herein, "heteroarylaliphatic (e.g., heteroaralkyl)" refers to an aliphatic group (e.g., C) substituted with a heteroaryl group_1-4Alkyl groups). "aliphatic", "alkyl" and "heteroaryl" have been defined above.

As used herein, "heteroaralkyl" refers to an alkyl group substituted with a heteroaryl group (e.g., C)_1-4Alkyl groups). Both "alkyl" and "heteroaryl" have been defined above. Heteroaralkyl is optionally substituted with one or more substituents, such as alkyl (including carboxyalkyl, hydroxyalkyl and haloalkyl, e.g., trifluoromethyl), alkenyl, alkynyl, cycloalkyl, (cycloalkyl) alkyl, heterocycloalkyl, (heterocycloalkyl) alkyl, aryl, heteroaryl, alkoxy, cycloalkoxy, heterocycloalkoxy, aryloxy, heteroaryloxy, aralkoxy, heteroaralkoxy, aroyl, heteroaroyl, nitro, carboxy, alkoxycarbonyl, alkylcarbonyloxy, aminocarbonyl, alkylcarbonylamino, cycloalkylcarbonylamino, (cycloalkylalkyl) carbonylamino, arylcarbonylamino, aralkylcarbonylamino, (heterocycloalkyl) carbonylamino, heteroarylcarbonylamino, heteroaralylcarbonylamino, cyano, halo, hydroxy, acyl, mercapto, alkylthioalkyl, cyano, hydroxy, acyl, mercapto, alkylthioalkyl, and the like, Sulfoxy, urea, thiourea, sulfamoyl, sulfonylamino, oxo, or carbamoyl.

As used herein, "acyl" refers to formyl or R^X-C (O) - (e.g. alkyl)-C (O) -, also known as "alkylcarbonyl"), wherein R^XAnd "alkyl" have been defined as before. Acetyl and pivaloyl are examples of acyl groups.

As used herein, "aroyl" or "heteroaroyl" refers to aryl-C (O) -or heteroaryl-C (O) -. The aryl and heteroaryl portions of the aroyl or heteroaroyl groups may be optionally substituted as defined hereinbefore.

"alkoxy" as used herein denotes alkyl-O-wherein "alkyl" has been defined as above.

As used herein, "carbamoyl" refers to a compound having the structure-O-CO-NR^XR^Yor-NR^X-CO-O-R^ZWherein R is^XAnd R^YHas been defined as above, R^ZCan be aliphatic, aryl, araliphatic, heterocycloaliphatic, heteroaryl or heteroarylaliphatic.

As used herein, "carboxyl" means-COOH, -COOR when used as a terminal group^X、-OC(O)H、-OC(O)R^X(ii) a or-OC (O) -or-C (O) O-when used as an internal group.

"Haloaliphatic" as used herein means an aliphatic group substituted with 1 to 3 halogens. For example, the term haloalkyl includes the group-CF₃。

"mercapto" as used herein means-SH.

As used herein, "sulfo" means-SO when used at the terminus₃H or-SO₃R^Xor-S (O) when used internally₃-。

As used herein, a "sulfonylamino" group refers to the structure-NR when used at the terminus^X-S(O)₂-NR^YR^ZAnd structure-NR when used internally^X-S(O)₂-NR^Y-, wherein R^X、R^YAnd R^ZAs already defined above.

As used herein, "sulfonamide" group meansStructure for terminal-S (O)₂-NR^XR^Yor-NR^X-S(O)₂-R²(ii) a Or structure-S (O) when used internally₂-NR^X-or-NR^X-S(O)₂-, wherein R^X、R^YAnd R^ZIs as defined above.

As used herein, "sulfanyl" refers to-S-R when used at the terminus^Xand-S-when used internally, wherein R^XAs already defined above. Examples of sulfanyl groups include aliphatic-S-, cycloaliphatic-S-, aryl-S-, and the like.

"sulfinyl" as used herein means-S (O) -R when used at the terminus^Xand-S (O) -when used internally, wherein R^XAs already defined above. Exemplary sulfinyl groups include aliphatic-S (O) -, aryl-S (O) -, (cycloaliphatic (aliphatic)) -S (O) -, cycloalkyl-S (O) -, heterocycloaliphatic-S (O) -, heteroaryl-S (O) -, and the like.

"Sulfonyl" as used herein means-S (O) when used for a terminus₂-R^XAnd when used internally-S (O)₂-, wherein R^XAs already defined above. Exemplary sulfonyl groups include aliphatic groups-S (O)₂-, aryl-S (O)₂-, (cycloaliphatic (aliphatic) group) -S (O)₂-, cycloaliphatic radical-S (O)₂-, heterocycloaliphatic-S (O)₂-, heteroaryl-S (O)₂-, (cycloaliphatic (amido (aliphatic))) -S (O)₂-and the like.

As used herein, "sulfoxy" means-O-SO-R when used at the terminus^Xor-SO-O-R^XAnd for internal use, -O-S (O) -or-S (O) -O-, wherein R^XAs already defined above.

"halogen" or "halogen" as used herein means fluorine, chlorine, bromine or iodine.

As used herein, "alkoxycarbonyl" is encompassed by the term carboxy, alone or in combination with another group, and is intended to denote a group such as alkyl-O-C (O) -.

"alkoxyalkyl" as used herein, means an alkyl group, such as alkyl-O-alkyl, wherein alkyl has been defined as above.

As used herein, "carbonyl" means-C (O) -.

As used herein, "oxo" refers to ═ O.

As used herein, "aminoalkyl" refers to the structure (R)^X)₂N-alkyl-.

"cyanoalkyl" as used herein means the structure (NC) -alkyl-.

As used herein, the "urea" group represents the structure-NR^X-CO-NR^YR^ZThe "thiourea" group represents the structure-NR when used at the terminal^X-CS-NR^YR^ZAnd structure-NR when used internally^X-CO-NR^Y-or-NR^X-CS-NR^Y-, wherein R^X、R^YAnd R^ZAs already defined above.

As used herein, the "guanidine" group represents the structure-N ═ C (N (R)^XR^Y))N(R^XR^Y) or-NR^X-C(＝NR^X)NR^XR^YWherein R is^XAnd R^YAs already defined above.

The term "amidino" as used herein denotes the structure-C ═ (NR)^X)N(R^XR^Y) Wherein R is^XAnd R^YAs already defined above.

In general, the term "ortho" refers to a substituent position on a group comprising two or more carbon atoms, wherein the substituent is attached to an adjacent carbon atom.

In general, the term "geminal" refers to a substituent position on a group comprising two or more carbon atoms, wherein the substituents are attached to the same carbon atom.

The terms "terminal" and "internal" refer to the position of a group within a substituent. When in useA group is terminal when it is present at the end of a substituent that is not further bonded to the remainder of the chemical structure. Carboxyalkyl, i.e. R^XO (O) C-alkyl is an example of a carboxyl group for the terminal. A group is internal when it is present in the middle of a substituent that is bonded to the rest of the chemical structure. Alkylcarboxy (e.g., alkyl-C (O) O-or alkyl-OC (O) -) and alkylcarboxylaryl (e.g., alkyl-C (O) O-aryl-or alkyl-O (CO) -aryl-) are examples of carboxyl groups for the interior.

As used herein, "cyclic group" includes mono-, di-and tri-cyclic ring systems including cycloaliphatic, heterocycloaliphatic, aryl or heteroaryl groups, each of which has been defined above.

As used herein, a "bridged bicyclic ring system" means a bicyclic heterocycloaliphatic ring system or a bicyclic cycloaliphatic ring system, wherein the rings are bridged. Examples of bridged bicyclic ring systems include, but are not limited to, adamantyl, norbornyl, bicyclo [3.2.1]Octyl, bicyclo [2.2.2]Octyl, bicyclo [3.3.1]Nonyl, bicyclo [3.2.3]Nonyl, 2-oxabicyclo [2.2.2]Octyl, 1-azabicyclo [2.2.2]Octyl, 3-azabicyclo [3.2.1]Octyl and 2, 6-dioxa-tricyclo [3.3.1.0^3，7]Nonyl radical. The bridged bicyclic ring system may optionally be substituted with one or more substituents, such as alkyl (including carboxyalkyl, hydroxyalkyl and haloalkyl, e.g., trifluoromethyl), alkenyl, alkynyl, cycloalkyl, (cycloalkyl) alkyl, heterocycloalkyl, (heterocycloalkyl) alkyl, aryl, heteroaryl, alkoxy, cycloalkoxy, heterocycloalkoxy, aryloxy, heteroaryloxy, aralkoxy, heteroaralkoxy, aroyl, heteroaroyl, nitro, carboxyl, alkoxycarbonyl, alkylcarbonyloxy, aminocarbonyl, alkylcarbonylamino, cycloalkylcarbonylamino, (cycloalkylalkyl) carbonylamino, arylcarbonylamino, aralkylcarbonylamino, (heterocycloalkyl) carbonylamino, heteroarylcarbonylamino, heteroaralkylcarbonylamino, cyano, halogen, hydroxy, acyl, mercapto, etc, Alkylsulfanyl, sulfoxy, urea, thiourea, sulfamoyl, sulfonylamino, oxo, or carbamoyl.

As used herein, "aliphatic chain" means a branched or straight chain aliphatic group (e.g., alkyl, alkenyl, or alkynyl). The linear aliphatic chain having the structure- [ CH ]₂]_v-, where v is 1 to 6. A branched aliphatic chain is a straight aliphatic chain substituted with one or more aliphatic groups. The branched aliphatic chain has the structure- [ CHQ ]]_v-, wherein Q is hydrogen or an aliphatic group; however, Q should in at least one instance be an aliphatic group. The term aliphatic chain includes alkyl, alkenyl and alkynyl chains, wherein alkyl, alkenyl and alkynyl are as defined above.

The phrase "optionally substituted" is used interchangeably with the phrase "substituted or unsubstituted". As described herein, the compounds of the invention may be optionally substituted with one or more substituents, such as those set forth generally above, or as exemplified by particular classes, subclasses, and species of the invention. The variable R is as described herein₁、R₂And R₃And other variables encompassed by formula I encompass specific groups such as alkyl and aryl groups. Unless otherwise noted, each particular group with respect to the variables Ra, Rb, Rc, Rd and other variables contained therein may be optionally substituted with one or more substituents described herein. Each substituent of a particular group is further optionally substituted with one to three of halo, cyano, oxo, alkoxy, hydroxy, amino, nitro, aryl, haloalkyl, and alkyl. For example, the alkyl group may be substituted with an alkylsulfanyl group, which may optionally be substituted with one to three of halogen, cyano, oxo, alkoxy, hydroxy, amino, nitro, aryl, haloalkyl, and alkyl. As additional examples, the cycloalkyl moiety of the (cycloalkyl) carbonylamino group can be optionally substituted with one to three of halo, cyano, alkoxy, hydroxy, nitro, haloalkyl, and alkyl. When two alkoxy groups are bonded to the same atom or adjacent atoms, the two alkoxy groups may form a ring together with the atom to which they are bonded.

In general, the term "substituted," whether preceded by the term "optionally," means that a hydrogen radical in a given structure is replaced with a radical that is designated as a substituent. Specific substituents are as defined above and described in the description of the compounds below and in the examples thereof. Unless otherwise specified, an optionally substituted group may have a substituent at each substitutable position of the group, and if more than one position in any given structure may be substituted with more than one substituent selected from a specified group, the substituents may be the same or different at each position. A ring substituent, such as heterocycloalkyl, may be bonded to another ring, such as cycloalkyl, to form a spiro-bicyclic ring system, e.g., the two rings share a common atom. As will be appreciated by those of ordinary skill in the art, the combinations of substituents contemplated by the present invention are those that form stable or chemically feasible compounds.

The phrase "stable or chemically feasible" as used herein means that the compounds are substantially unchanged when subjected to the conditions used for their preparation, detection, preferably recovery, purification, and for one or more of the purposes disclosed herein. In some embodiments, a stable compound or chemically feasible compound is one that remains substantially unchanged in the absence of moisture or other chemically reactive conditions at a temperature of 40 ℃ or less for at least one week.

An effective amount, as used herein, is defined as the amount required to confer a therapeutic effect on the patient being treated, and is generally determined based on the age, body surface area, weight and condition of the patient. Animal-to-human dose correlations (based on milligrams per square meter of body surface) as described by Freireich et al, Cancer chemither. 219 (1966). The body surface area can be determined approximately from the height and weight of the patient. See, for example, scientific tables, Geigy Pharmaceuticals, Ardsley, New York, 537 (1970). As used herein, "patient" means a mammal, including a human.

Unless otherwise specified, structures depicted herein are also meant to include all isomeric (e.g., enantiomeric, diastereomeric, and geometric (or conformational) forms of the structure; for example, the R and S configuration of each asymmetric center, (Z) and(E) double bond isomers, and conformational isomers of (Z) and (E). Thus, single stereochemical isomers as well as enantiomeric, diastereomeric, and geometric (or conformational) mixtures of these compounds are within the scope of the invention. Unless otherwise specified, all tautomeric forms of the compounds of the invention are within the scope of the invention. In addition, unless otherwise specified, the structures depicted herein are also meant to include compounds that differ only in the presence of one or more isotopically enriched atoms. For example, except that hydrogen is replaced by deuterium or tritium or carbon is replaced by¹³C-or¹⁴C-enriched carbon instead of compounds having the structure of the present invention are within the scope of the present invention. Such compounds are useful, for example, as analytical tools or probes in biological assays.

II. Compound

Compounds useful for modulating ABC transporter and CFTR activity have the general structure of formula I:

or a pharmaceutically acceptable salt thereof, wherein:

each Ra is independently H, an optionally substituted aliphatic, an optionally substituted aryl, an optionally substituted aralkyl, an optionally substituted heteroaralkyl, an optionally substituted heteroaryl, an optionally substituted cycloaliphatic, or an optionally substituted cycloheteroaliphatic.

Each Rb is independently an optionally substituted aliphatic, an optionally substituted aryl, an optionally substituted aralkyl, an optionally substituted heteroaralkyl, an optionally substituted heteroaryl, an optionally substituted cycloaliphatic, an optionally substituted cycloheteroaliphatic, an optionally substituted heteroaryl, a,Wherein w is 1, 2, 3, 4 or 5, phenylOptionally substituted with 1-4 Re, or Ra and Rb together with the nitrogen atom to which they are bound form an optionally substituted heterocycloaliphatic, optionally substituted heteroaryl.

Each Rc is independently H, an optionally substituted heterocycloaliphatic, an optionally substituted cycloaliphatic, or an unsubstituted aliphatic.

Each Rd is independently H, an optionally substituted aliphatic group, an optionally substituted aryl group, an optionally substituted aralkyl group, an optionally substituted heteroaralkyl group, an optionally substituted heteroaryl group, an optionally substituted cycloaliphatic group, an optionally substituted cycloheteroaliphatic group, or Rc and Rd together with the nitrogen atom to which they are bound form an optionally substituted heterocycloaliphatic group.

Ring a is aryl or heteroaryl, each optionally substituted with 1-4 Re.

Each Re is independently carboxy, amino, nitro, alkylcarbonylamino, cycloalkylcarbonylamino, (cycloalkyl) alkylcarbonylamino, arylcarbonylamino, aralkylcarbonylamino, (heterocycloalkyl) carbonylamino, (heterocycloalkyl) alkylcarbonylamino, heteroarylcarbonylamino, heteroaralkylcarbonylamino, sulfonyl, sulfinyl, sulfanyl, sulfoxy, sulfamoyl, sulfonylamino, ketal, carbamoyl, cyano, halogen, urea, thiourea, haloalkyl, or-Z-Rf, wherein each alkyl, cycloalkyl, heterocycloalkyl, aryl, and heteroaryl portion of the Re substituent is optionally substituted with 1-3 Rg, or two Re's on adjacent a ring atoms together with the a ring atom to which they are bonded form a heterocycloaliphatic ring.

Each Z is absent, -O-or-S-.

Each Rf is independently hydrogen, alkyl, carboxyalkyl, hydroxyalkyl, alkenyl, alkynyl, cycloalkyl, (cycloalkyl) alkyl, heterocycloalkyl, (heterocycloalkyl) alkyl, aryl, heteroaryl, aralkyl, heteroaralkyl, aroyl, heteroaroyl, alkoxycarbonyl, alkylcarbonyl, aminocarbonyl, or acyl, wherein each alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl, and heteroaryl moiety on the Rf substituent is optionally substituted with 1-3 Rg.

Each Rg is independently halogen, cyano, alkoxy, hydroxy, nitro, haloalkyl, and alkyl.

However, in several embodiments, when Rd is alkyl substituted with 4-methyl-1-piperazinyl, then Ring A is not 3, 4, 5-trimethoxyphenyl; ring a is not 7-chloro-2- (4-fluorophenyl) pyrazolo [1, 5-a ] pyridin-3-yl or 7-cyclopentylamino-2- (4-fluorophenyl) pyrazolo [1, 5-a ] pyridin-3-yl; when Rc is H, Ra and Rb are both alkyl, and ring a is oxazolyl, then Rd is not {1- [ (3, 5-difluorophenyl) methyl) ] -2-hydroxy-4- (1H-pyrazol-3-yl) butyl).

A subset of compounds useful for modulating ABC transporter and CFTR activity have the structure of formula II:

wherein the variables Ra, Rb, Rc, Rd and Re are as defined above, with the proviso that when Rd is alkyl substituted by 4-methyl-1-piperazinyl then ring A is not 3, 4, 5-trimethoxyphenyl.

Another group of compounds useful for modulating ABC transporter and CFTR activity have the structure of formula III:

wherein ring B is an optionally substituted heterocycloaliphatic and the variables Rc, Rd and Re are as defined above.

Embodiments of the compounds of formulae I, II and III include the following.

A. Ring A

Different aspects of ring a include the following:

ring a is an aryl group, such as phenyl. Ring a is a bicyclic aryl group, such as naphthyl and azulenyl. In a particular aspect, ring a is naphthyl. Embodiments of aspects wherein ring a is aryl include the following. Ring a is aryl optionally substituted with 1-4 Re. Ring a is aryl substituted with at least one Re substituent. Ring a is an aryl group substituted with at least one Re substituent ortho or meta to the point of attachment between ring a and the pyrimidine. Ring a is an aryl group substituted with at least one Re substituent ortho, para, or meta to the point of attachment between ring a and the pyrimidine. Ring a is an aryl group substituted ortho to the point of attachment between ring a and the pyrimidine with at least one Re substituent. Ring a is aryl substituted with at least one Re substituent para to the point of attachment between ring a and the pyrimidine. Ring a is aryl substituted with at least one Re substituent meta to the point of attachment between ring a and the pyrimidine. Ring a is an aryl group including at least two Re substituents. Ring a is an aryl group including at least two Re substituents, one of which is ortho to the point of attachment between ring a and the pyrimidine.

Ring a is heteroaryl. Ring a is a monocyclic heteroaryl group, such as pyrrolyl, furyl, oxazolyl, thiazolyl, pyrazolyl, thienyl, pyridazinyl, pyrazinyl, pyrimidinyl or pyridyl. Ring a is thienyl, furyl, pyrimidinyl, or pyridyl. Ring a is thienyl. Ring A is a bicyclic heteroaryl group such as indolizinyl, indolyl, isoindolyl, indazolyl, benzimidazolyl, purinyl, isoquinolyl, benzofuranyl, quinolinyl, benzothienyl, or benzodioxolanyl. Ring A is benzothienyl, benzodioxolanyl. Ring a is heteroaryl optionally substituted with 1-4 Re. Ring a is heteroaryl substituted with at least one Re substituent. Ring a is heteroaryl substituted with at least one Re substituent ortho, para, or meta to the point of attachment between ring a and the pyrimidine. Ring a is heteroaryl substituted ortho to the point of attachment between ring a and the pyrimidine with at least one Re substituent. Ring a is heteroaryl substituted with at least one Re substituent para to the point of attachment between ring a and the pyrimidine. Ring a is heteroaryl substituted with at least one Re substituent meta to the point of attachment between ring a and the pyrimidine. Ring a is heteroaryl including at least two Re substituents. Ring a is a heteroaryl group including at least two Re substituents, one of which is ortho to the point of attachment between ring a and the pyrimidine.

B. Substituent Re

Various aspects of the Re substituent include the following:

each Re substituent, if present, is independently carboxy, amino, nitro, alkylcarbonylamino, cycloalkylcarbonylamino, (cycloalkyl) alkylcarbonylamino, arylcarbonylamino, aralkylcarbonylamino, (heterocycloalkyl) carbonylamino, (heterocycloalkyl) alkylcarbonylamino, heteroarylcarbonylamino, heteroaralkylcarbonylamino, sulfonyl, sulfinyl, sulfanyl, sulfoxy, sulfamoyl, sulfonylamino, ketal, or carbamoyl, cyano, halogen, urea, thiourea, haloalkyl, wherein each alkyl, cycloalkyl, heterocycloalkyl, aryl, and heteroaryl portion of the Re substituent is optionally substituted with 1-3 of halogen, cyano, alkoxy, hydroxy, nitro, haloalkyl, and alkyl.

The Re substituents, if present, are each independently carboxy, nitro, cyano, halo, haloalkyl, wherein each alkyl, cycloalkyl, heterocycloalkyl, aryl, and heteroaryl portion of the Re substituent is optionally substituted with 1-3 of halo, cyano, alkoxy, hydroxy, nitro, haloalkyl, and alkyl.

The Re substituents, if present, are each independently amino, alkylcarbonylamino, cycloalkylcarbonylamino, (cycloalkyl) alkylcarbonylamino, arylcarbonylamino, aralkylcarbonylamino, (heterocycloalkyl) carbonylamino, (heterocycloalkyl) alkylcarbonylamino, heteroarylcarbonylamino, heteroaralkylcarbonylamino, urea, thiourea, wherein each alkyl, cycloalkyl, heterocycloalkyl, aryl and heteroaryl portion of the Re substituent is optionally substituted with 1-3 halo, cyano, alkoxy, hydroxy, nitro, haloalkyl and alkyl groups.

The Re substituents, if present, are each independently alkylsulfanyl, sulfoxyloxy, sulfamoyl, sulfonylamino, ketal, or carbamoyl, wherein each alkyl, cycloalkyl, heterocycloalkyl, aryl, and heteroaryl portion of the Re substituent is optionally substituted with 1-3 of halo, cyano, alkoxy, hydroxy, nitro, haloalkyl, and alkyl.

The Re substituent, if present, is independently-Z-Rf, wherein each Z is absent, -O-or-S-, each Rf is independently hydrogen, alkyl, carboxyalkyl, hydroxyalkyl, alkenyl, alkynyl, cycloalkyl, (cycloalkyl) alkyl, heterocycloalkyl, (heterocycloalkyl) alkyl, aryl, heteroaryl, aralkyl, heteroaralkyl, aroyl, heteroaroyl, alkoxycarbonyl, alkylcarbonyl, aminocarbonyl, or acyl, wherein each alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl, and heteroaryl portion of the Rf substituent is optionally substituted with 1-3 of halo, cyano, alkoxy, hydroxy, nitro, haloalkyl, and alkyl.

The Re substituent, if present, is independently-Z-Rf, wherein Z is absent, and each Rf is independently hydrogen, alkyl, carboxyalkyl, hydroxyalkyl, alkenyl, alkynyl, (cycloalkyl) alkyl, (heterocycloalkyl) alkyl, aralkyl, heteroaralkyl, aroyl, heteroaroyl, wherein each alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl, and heteroaryl moiety on the Rf substituent is optionally substituted with 1-3 halo, cyano, alkoxy, hydroxy, nitro, haloalkyl, and alkyl groups.

The Re substituent, if present, is independently-Z-Rf, wherein each Z is absent, each Rf is independently cycloalkyl, heterocycloalkyl, aryl, heteroaryl, alkoxycarbonyl, alkylcarbonyl, aminocarbonyl, or acyl, wherein each alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl, and heteroaryl moiety on the Rf substituent is optionally substituted with 1-3 halo, cyano, alkoxy, hydroxy, nitro, haloalkyl, and alkyl groups.

The Re substituent, if present, is independently alkyl, hydroxyalkyl, haloalkyl, haloalkoxy, alkylOxy, halogen, hydroxy, alkoxycarbonyl, alkylcarbonylamino, aryloxy, sulfoxy, carboxy, acyl or alkylcarbonyl. Re substituents of ring A are independently chlorine, bromine, phenoxy, -CF₃Alkoxy (e.g., methoxy, ethoxy), -C (O) Oalkyl (e.g., -C (O) O-methyl, -C (O) O-ethyl, -C (O) O-propyl, and-C (O) O-isopropyl), alkyl (e.g., methyl, ethyl, propyl, butyl, and isopropyl), haloalkoxy (e.g., CF₃O-), hydroxyalkyl (e.g., HO-methyl-, HO-ethyl-, and HO-propyl), alkylcarbonylamino (e.g., methyl-C (O) -NH-, ethyl-C (O) -NH-, propyl-C (O) -NH-, and isopropyl-C (O) -NH-), -S (O)₂Alkyl (e.g., -S (O)₂Methyl, -S (O)₂Ethyl, -S (O)₂Butyl and-S (O)₂Propyl) and-S (O)₂And (4) an aryl group.

Two Re substituents, if present on adjacent a ring atoms, together with the ring atoms to which they are bonded form a heterocycloaliphatic (e.g., a dioxolane ring).

C. Substituents Ra and Rb

Different aspects of Ra include the following:

ra is hydroxyalkyl, alkoxyalkyl, (heterocycloalkyl) alkyl, (cycloalkyl) alkyl, alkoxycarbonylalkyl, aminoalkyl, carboxyalkyl, (cycloalkoxy) alkyl, (heterocycloalkoxy) alkyl, (aryloxy) alkyl, (heteroaryloxy) alkyl, (aralkoxy) alkyl, (heteroarylalkoxy) alkyl, (aminocarbonyl) alkyl, (alkylcarbonylamino) alkyl, (cycloalkylcarbonylamino) alkyl, (cycloalkyl-alkylcarbonylamino) alkyl, (arylcarbonylamino) alkyl, (aralkylcarbonylamino) alkyl, (heterocycloalkyl-carbonylamino) alkyl, (heterocycloalkyl-alkylcarbonylamino) alkyl, (heteroarylcarbonylamino) alkyl, (heteroaralkylcarbonylamino) alkyl, (urea) alkyl, (thiourea) alkyl, (sulfamoyl) alkyl, cycloalkyl-carbonylamino, and the like, (sulfonylamino) alkyl, (alkoxycarbonyl) alkyl or (alkylcarbonyloxy) alkyl.

Ra is H.

Ra is an optionally substituted aliphatic group, such as an optionally substituted alkyl group, an optionally substituted alkenyl group, or an optionally substituted alkynyl group. Ra is an optionally substituted alkenyl. Ra is alkenyl. Ra is propenyl, butenyl or pentenyl. Ra is alkenyl substituted with one or more hydroxy, amino, halogen, cyano, oxo. Ra is an optionally substituted alkyl group. Ra is an optionally substituted methyl group, an optionally substituted ethyl group, an optionally substituted propyl group, an optionally substituted butyl group and an optionally substituted pentyl group. Ra is alkyl substituted with one or more hydroxy, alkoxy, amino, halo, cyano, oxo. Ra is methyl substituted with one or more hydroxy, alkoxy, amino, halo, cyano, oxo. Ra is ethyl substituted with one or more hydroxy, alkoxy, amino, halo, cyano, oxo. Ra is propyl substituted with one or more hydroxy, alkoxy, amino, halo, cyano, oxo. Ra is butyl substituted with one or more hydroxy, alkoxy, amino, halo, cyano, oxo. Ra is hydroxyalkyl (e.g., HO-methyl, HO-ethyl, HO-butyl, and HO-propyl). Ra is alkoxyalkyl (e.g. methoxymethyl-, methoxyethyl-, methoxypropyl-, methoxybutyl-, ethoxymethyl-, ethoxyethyl-, ethoxypropyl-, ethoxybutyl-, propoxymethyl-, propoxyethyl-and propoxypropyl-. Ra is cyanoalkyl (e.g. NC-methyl, NC-ethyl, NC-propyl and NC-butyl). Ra is carboxyalkyl (e.g. methyl-O (O) C-methyl-, methyl-O (O) C-ethyl-, methyl-O (O) C-propyl-, methyl-O (O) C-butyl-, ethyl-O (O) C-methyl-, ethyl-O (O) C-ethyl-), ethyl-O (O) C-propyl, ethyl-O (O) C-butyl, propyl-O (O) C-methyl-, propyl-O (O) C-ethyl and propyl-O (O) C-propyl).

Ra is an optionally substituted aryl, optionally substituted aralkyl, optionally substituted heteroaralkyl, optionally substituted heteroaryl, optionally substituted cycloaliphatic, or optionally substituted cycloheteroaliphatic. Ra is an optionally substituted aralkyl group (e.g., an optionally substituted benzyl group and an optionally substituted phenethyl group). Ra is optionally substituted heteroaralkyl (e.g., optionally substituted furanylmethyl, optionally substituted pyridylmethyl, and optionally substituted pyridylethyl).

Different aspects of Rb include the following:

rb is an optionally substituted aliphatic group such as an optionally substituted alkyl group and an optionally substituted alkenyl group or an optionally substituted alkynyl group. Rb is an optionally substituted alkenyl. Rb is alkenyl. Rb is propenyl, butenyl or pentenyl. Rb is alkenyl substituted with one or more of hydroxy, amino, halo, cyano, and oxo. Rb is an optionally substituted alkyl. Rb is optionally substituted methyl, optionally substituted ethyl, optionally substituted propyl, optionally substituted butyl and optionally substituted pentyl. Rb is alkyl substituted with one or more hydroxy, alkoxy, amino, halo, cyano, oxo. Rb is methyl substituted with one or more hydroxy, alkoxy, amino, halo, cyano, and oxo. Rb is ethyl substituted with one or more hydroxy, alkoxy, amino, halo, cyano, oxo. Rb is propyl substituted with one or more hydroxy, alkoxy, amino, halo, cyano, oxo. Rb is butyl substituted with one or more hydroxy, alkoxy, amino, halo, cyano, oxo. Rb is hydroxyalkyl (e.g., HO-methyl, HO-ethyl, HO-butyl, and HO-propyl). Rb is alkoxyalkyl (e.g. methoxymethyl-, methoxyethyl-, methoxypropyl-, methoxybutyl-, ethoxymethyl-, ethoxyethyl-, ethoxypropyl-, ethoxybutyl-, propoxymethyl-, propoxyethyl-and propoxypropyl-. Rb is cyanoalkyl (e.g. NC-methyl, NC-ethyl, NC-propyl and NC-butyl). Rb is carboxyalkyl (e.g. methyl-O (O) C-methyl-, methyl-O (O) C-ethyl-, methyl-O (O) C-propyl-, methyl-O (O) C-butyl-, ethyl-O (O) C-methyl-, ethyl-O (O) C-ethyl-), ethyl-O (O) C-propyl, ethyl-O (O) C-butyl, propyl-O (O) C-methyl-, propyl-O (O) C-ethyl and propyl-O (O) C-propyl).

Rb is an optionally substituted aryl, an optionally substituted aralkyl, an optionally substituted heteroaralkyl, an optionally substituted heteroaryl, an optionally substituted cycloaliphatic or an optionally substituted cycloheteroaliphatic. Rb is an optionally substituted aralkyl group (e.g., an optionally substituted benzyl group and an optionally substituted phenethyl group). Rb is an optionally substituted heteroaralkyl (e.g., an optionally substituted furanylmethyl, an optionally substituted pyridylmethyl, and an optionally substituted pyridylethyl).

Rb isWherein w is 1, 2, 3, 4 or 5, phenyl is optionally substituted with 1-4 Re. Rb isWherein w is 1 and phenyl is optionally substituted with 1-4 Re. Rb isWherein w is 3 and phenyl is optionally substituted with 1-4 Re.

Different aspects of Ra and Rb include the following:

ra and Rb together with the nitrogen atom to which they are bound form an optionally substituted heterocycloaliphatic or an optionally substituted heteroaryl. Ra and Rb together with the nitrogen atom to which they are bound form an optionally substituted heterocycloaliphatic group (e.g., an optionally substituted azocane, an optionally substituted aza * group, an optionally substituted piperidine, an optionally substituted pyrrolidine, an optionally substituted tetrahydropyridine, an optionally substituted pyrroline, an optionally substituted piperazine, an optionally substituted azetidine, an optionally substituted morpholino, an optionally substituted thiomorpholino, an optionally substituted perhydroquinoline, an optionally substituted tetrahydroisoquinoline, and an optionally substituted thiapyrrolidine). Ra and Rb together with the nitrogen atom to which they are bound form an optionally substituted 8-membered heterocycloaliphatic. Ra and Rb together with the nitrogen atom to which they are bound form an optionally substituted 7-membered heterocycloaliphatic. Ra and Rb together with the nitrogen atom to which they are bound form an optionally substituted 6-membered heterocycloaliphatic. Ra and Rb taken together with the nitrogen atom to which they are bondedAn optionally substituted 5-membered heterocycloaliphatic. Ra and Rb together with the nitrogen atom to which they are bound form a heterocycloaliphatic substituted with 1-3 halogens, haloalkyl groups, alkyl groups, alkoxycarbonyl groups, alkylcarbonyl groups, aminocarbonyl groups, hydroxyalkyl groups, sulfoxy groups, sulfanyl groups, sulfonyl groups, sulfinyl groups, optionally substituted aryl groups, optionally substituted alkoxy groups, optionally substituted aralkyl groups, optionally substituted aroyl groups, optionally substituted heterocycloaliphatic groups, or optionally substituted heteroaryl groups. Ra and Rb together with the nitrogen atom to which they are bonded form a substituted group consisting of 1-3 aryl (e.g., phenyl), acetyl, aralkyl (e.g., benzyl), hydroxyalkyl (e.g., HO-methyl and HO-ethyl), heterocycloaliphatic (e.g., dioxolanyl or 2-oxo-1, 3-dihydrobenzimidazol-1-yl), optionally substituted heteroaryl (e.g., 2-oxo-benzimidazolyl), phenyl-C (O) -, 4-halo-phenyl-C (O) -, optionally substituted phenyl (e.g., 4-cyanophenyl-, 4-hydroxyphenyl), and phenyl S (O)₂-a substituted heterocycloaliphatic group.

Ra and Rb together with the nitrogen atom to which they are bound form an optionally substituted heteroaryl group (e.g., imidazoline, pyrrole, pyrazole, tetrahydroquinoline).

Ra and Rb are both optionally substituted aliphatic groups. Ra and Rb are both optionally substituted alkyl. Ra and Rb are both alkyl groups. Ra and Rb are both ethyl.

In some embodiments, Ra and Rb are both H.

D. Substituents Rc and Rd

Different aspects of Rd include the following:

rd is H or an optionally substituted aliphatic group. And Rd is H. Rd is optionally substituted alkyl (e.g., optionally substituted methyl, optionally substituted ethyl, optionally substituted propyl, and optionally substituted butyl). Rd is methyl, ethyl, propyl, isopropyl or butyl.

Rd is optionally substituted aralkyl (e.g., optionally substituted benzyl and optionally substituted phenethyl). Rd is optionally substituted heteroaralkyl (e.g., optionally substituted furanylmethyl, optionally substituted pyridylmethyl, and optionally substituted pyridylethyl). Rd is an optionally substituted heteroaryl (e.g., an optionally substituted pyridyl, an optionally substituted pyrrolyl, and an optionally substituted furanyl). Rd is optionally substituted aryl (e.g., optionally substituted phenyl). Rd is an optionally substituted cycloaliphatic group (e.g., an optionally substituted cyclopentyl group and an optionally substituted cyclohexyl group). Rd is an optionally substituted heterocycloaliphatic (e.g., an optionally substituted piperidine).

Different aspects of Rc include the following:

rc is H. Rc is an optionally substituted heterocycloaliphatic (e.g., an optionally substituted piperidine). Rc is an optionally substituted cycloaliphatic group (e.g., an optionally substituted cyclopentyl group and an optionally substituted cyclohexyl group). Rc is an unsubstituted aliphatic group. Rc is unsubstituted alkyl. Rc is methyl, ethyl, propyl or butyl.

Different aspects of Rc and Rd include the following:

rc and Rd together with the nitrogen atom to which they are bonded form an optionally substituted heterocycloaliphatic. Rc and Rd together with the nitrogen atom to which they are bound form optionally substituted pyrrolines, pyrrolidines, imidazolines, imidazolidines, piperidines, piperazines, tetrahydropyridines, morpholines, and thiomorpholines.

Rc and Rd are both H.

Specific compounds useful for modulating ABC transporter and/or CFTR activity are based on general formula I, and sub-formulae II and III may include different combinations of each of the aspects and embodiments described above. For example, a particular compound or subgroup of compounds may comprise different combinations of the above aspects.

In certain embodiments, compounds useful for modulating ABC transporter and/or CFTR activity have the structures listed in table 1.

Table 1: specific compounds

III. Synthesis method

Compounds of formulae I, II and III can be produced via known synthetic methods. Schemes 1 and 2 illustrate two possible methods for producing compounds of formula I.

Scheme 1:

a) EtONa, EtOH; b) EtONa, EtOH, thiourea. * H₂SO₄(ii) a c) PyBOP or EDC, NHRcRd; d) mCPBA or H₂O₂，CH₃CO₂H; HNRaRb, dioxane, 85 ℃.

In step a of scheme 1, a ketone is reacted with diethyl oxalacetate in the presence of a base (e.g., sodium ethoxide) and an organic solvent (e.g., ethanol) to provide dioxobutyric acid. In step b, the dioxobutyric acid is refluxed in the presence of a base (e.g., EtONa) and 2-methylisothiourea, followed by organic extraction and acid washing to give 2- (methylthio) pyrimidine-4-carboxylic acid. In step c, the carboxylic acid is converted to the amide using standard amine coupling procedures, for example with PyBOP or EDC in the presence of an appropriate amine (e.g. NHRcRd). In step d, in CH₃CO₂In H, thiols with oxidizing agents (e.g. m-CPBA or H)₂O₂) Oxidation to obtain sulfinyl or sulfonyl compound. In step eIn an organic solvent (e.g. dioxane), the sulfinyl or sulfonyl compound is reacted with the appropriate amine HNRaRb to give the compound of formula I.

And (2) a flow scheme:

a)NaOH，H₂O；b)i)SOCl₂PhMe is refluxed; ii) MeOH, DIEA, CHCl₃，0℃c)NHRcRd，CHCl₃；d)A-B(OR)₂，PdCl₂(dppf)，K₃PO₄，DMF/DME/H₂O，75℃，e)i)mCPBA，CHCl₃；ii)Me₂S；f)HNRaRb，NMM，DMSO，75℃。

In scheme 2, step a, diethyl oxalacetate is reacted with methylisothiourea to give 2-methylsulfanyl-6-oxo-1, 6-dihydro-pyrimidin-4-yl-carboxylic acid. In step b, the carboxylic acid is refluxed with thionyl chloride in an organic solvent (e.g. toluene) followed by treatment with methanol to give methyl 2-methylsulfanyl-6-chloro-pyrimidin-4-ylcarboxylate. In step c, the methyl ester is converted to the amide using an appropriate amine (e.g., NHRcRd). In step d, a coupling agent (e.g., PdCl) is used₂(dppf)) boronic acid derivatives of Ring A (e.g., [ Ring A ]]-B(OR)₂) Coupled with pyrimidine-4-carboxylic acid amides. The desired compounds are formed via synthesis steps e and f, which are analogous to steps d and e in scheme 1.

Use, route of administration and formulation

Thus, in another aspect of the invention, pharmaceutically acceptable compositions are provided, wherein these compositions comprise any of the compounds as described herein, and optionally a pharmaceutically acceptable carrier, adjuvant or vehicle. In certain embodiments, these compositions optionally further comprise one or more additional therapeutic agents.

It will also be appreciated that certain compounds of the invention can be present in free form for use in therapy, or optionally pharmaceutically acceptable derivatives thereof. According to the present invention, pharmaceutically acceptable derivatives include, but are not limited to, pharmaceutically acceptable salts, esters, salts of such esters, or any other adduct or derivative capable of providing, directly or indirectly, a compound as described herein or a metabolite or residue thereof upon administration to a patient in need thereof.

The term "pharmaceutically acceptable salt" as used herein, means those salts which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of human beings and lower animals without undue toxicity, irritation, allergic response and the like, commensurate with a reasonable benefit/risk ratio. "pharmaceutically acceptable salt" means any non-toxic salt or ester salt of a compound of the present invention which, upon administration to a recipient, is capable of providing, directly or indirectly, a compound of the present invention or an inhibitorily active metabolite or residue thereof.

Pharmaceutically acceptable salts are well known in the art. Pharmaceutically acceptable salts are described in detail, for example, in j.pharmaceutical Sciences, 1977, 66, 1-19, by s.m.berge et al, which is incorporated herein by reference. Pharmaceutically acceptable salts of the compounds of the present invention include those derived from suitable inorganic and organic acids and bases. Examples of pharmaceutically acceptable non-toxic acid addition salts are amino salts formed with inorganic acids such as hydrochloric acid, hydrobromic acid, phosphoric acid, sulfuric acid and perchloric acid, or with organic acids such as acetic acid, oxalic acid, maleic acid, tartaric acid, citric acid, succinic acid or malonic acid, or by other methods used in the art, such as ion exchange. Other pharmaceutically acceptable salts include adipates, alginates, ascorbates, aspartates, benzenesulfonates, benzoates, bisulfates, borates, butyrates, camphorates, camphorsulfonates, citrates, cyclopentanepropionates, digluconates, dodecylsulfates, ethanesulfonates, formates, fumarates, glucoheptanoates, glycerophosphates, gluconates, hemisulfates, heptanoates, hexanoates, hydroiodides, 2-hydroxyhydroxyglucarates, salts of adipic acid, salts of benzoic acid, salts of hydrogen sulfate, salts of benzoic acid, salts of butyric acid, salts of camphorsulfonic acidEthanesulfonates, lactobionates, lactates, laurates, malates, maleates, malonates, methanesulfonates, 2-naphthalenesulfonates, nicotinates, nitrates, oleates, oxalates, palmitates, embonates, pectinates, persulfates, 3-phenylpropionates, phosphates, picrates, pivalates, propionates, stearates, succinates, sulfates, tartrates, thiocyanates, p-toluenesulfonates, undecanoates, valerates, and the like. Salts derived from suitable bases include alkali metals, alkaline earth metals, ammonium and N⁺(C_1-4Alkyl radical)₄And (3) salt. The invention also encompasses quaternization of any basic nitrogen-containing group of the compounds as disclosed herein. By means of such quaternization, products which are soluble or dispersible in water or oil can be obtained. Representative alkali or alkaline earth metal salts include sodium, lithium, potassium, calcium, magnesium, and the like. Other pharmaceutically acceptable salts include, when appropriate, non-toxic ammonium, quaternary ammonium and amine cations, generated using counterions such as halides, hydroxides, carboxylates, sulfates, phosphates, nitrates, lower alkyl sulfonates and aryl sulfonates.

As noted above, the pharmaceutically acceptable compositions of the present invention additionally comprise a pharmaceutically acceptable carrier, adjuvant or vehicle, as described herein, including any and all solvents, diluents or other liquid excipients, dispersing or suspending aids, surfactants, isotonicity agents, thickening or emulsifying agents, preservatives, solid binders, lubricants and the like as appropriate for the particular dosage form desired. Remington's Pharmaceutical Sciences, SixteenthEdition, e.w. martin (Mack Publishing co., Easton, Pa., 1980) disclose various carriers for formulating pharmaceutically acceptable compositions and known techniques for their preparation. Except insofar as any conventional carrier medium is incompatible with the compounds of the invention, e.g., any other component that produces any undesirable biological effect or interacts in a deleterious manner with a pharmaceutically acceptable composition, its use is contemplated as falling within the scope of the present invention. Some examples of materials capable of serving as pharmaceutically acceptable carriers include, but are not limited to, ion exchangers; alumina; aluminum stearate; lecithin; serum proteins, such as human serum albumin; buffer substances, such as phosphates; glycine; sorbic acid or potassium sorbate; partial glyceride mixtures of saturated vegetable fatty acids; water; salts or electrolytes, such as protamine sulfate, disodium hydrogen phosphate, potassium hydrogen phosphate, sodium chloride, zinc salts; colloidal silicon dioxide; magnesium trisilicate; polyvinylpyrrolidone; a polyacrylate; waxes; polyethylene-polypropylene oxide-block polymers; lanolin; sugars such as lactose, glucose and sucrose; starches, such as corn starch and potato starch; cellulose and its derivatives, such as sodium carboxymethyl cellulose, ethyl cellulose and cellulose acetate; crushed tragacanth; malt; gelatin; talc; excipients, such as cocoa butter and suppository waxes; oils such as peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, corn oil and soybean oil; glycols, such as propylene glycol or polyethylene glycol; esters such as ethyl oleate and ethyl laurate; agar; buffering agents such as magnesium hydroxide and aluminum hydroxide; alginic acid; pyrogen-free water; isotonic saline; ringer's solution; ethanol; a phosphate buffer solution; and other non-toxic compatible lubricants such as sodium lauryl sulfate and magnesium stearate; coloring agents, releasing agents, coating agents, sweetening, flavoring and perfuming agents, preserving and anti-oxidizing agents may also be present in the composition, according to the judgment of the person skilled in the art.

In another aspect, the invention provides methods of treating a disorder, disease, or condition involving ABC transporter activity. In certain embodiments, the present invention provides methods of treating a disorder, disease, or condition involving a deficiency in ABC transporter activity, the method comprising administering to a subject, preferably a mammal, in need thereof a composition comprising a compound of formula (I).

In certain embodiments, the present invention provides methods of treating: cystic fibrosis, hereditary emphysema, hereditary hemochromatosis, coagulation-fibrinolysis deficiency (e.g., protein C deficiency), hereditary angioedema type 1, lipid processing deficiency (e.g., familial hypercholesterolemia), chylomicronemia type 1, β -lipoproteinemia, lysosomal storage diseases (e.g., I-cell disease/pseudo Hurler), mucopolysaccharidosis, Sandhof/Tay-Sachs, Crigler-Najjar type II, polyendocrinosis/hyperinsulinemia, diabetes, Laron dwar, myeloperoxidase deficiency, primary hypoparathyroidism, melanoma, polyoses type CDG 1, hereditary emphysema, congenital hyperthyroidism, osteogenesis imperfecta, hereditary hypofibrinogenemia, ACT deficiency, Diabetes Insipidus (DI), metaplasia (DI), nephrogenic DI, Charcot-Marie syndrome, Marian-Marie syndrome, Marie-Mar, -Behcet's disease, neurodegenerative diseases (e.g. Alzheimer's disease, Parkinson's disease, amyotrophic lateral sclerosis, progressive supranuclear palsy, pick's disease), several polyglutamine neurological disorders (e.g. Huntington's disease, spinocerebellar ataxia type I, spinal and bulbar muscular atrophy, dentatorubral-pallidoluysian and myotonic dystrophy) and spongiform encephalopathies (e.g. hereditary Creutzfeldt-Jakob disease (caused by prion protein processing defects), Fabry's disease and Stewart-Schneider syndrome), secretory diarrhoea, polycystic kidney disease, Chronic Obstructive Pulmonary Disease (COPD), dry eye disease and sjogren's disease comprising the step of administering to said mammal an effective amount of a composition comprising a compound of formula (I) or the preferred embodiments thereof as described above.

According to an alternative preferred embodiment, the present invention provides a method of treating cystic fibrosis comprising the step of administering to said mammal an effective amount of a composition comprising a compound of formula (I) or a preferred embodiment thereof as described above.

An "effective amount" of a compound or pharmaceutically acceptable composition according to the present invention is an amount effective to treat or reduce the severity of one or more of the following diseases: cystic fibrosis, hereditary emphysema, hereditary hemochromatosis, coagulation-fibrinolysis deficiency (e.g., protein C deficiency), hereditary angioedema type 1, lipid processing deficiency (e.g., familial hypercholesterolemia), chylomicronemia type 1, β -lipoproteinemia, lysosomal storage diseases (e.g., I-cell disease/pseudo Hurler), mucopolysaccharidosis, Sandhof/Tay-Sachs, Crigler-Najjar type II, polyendocrinosis/hyperinsulinemia, diabetes, Laron dwar, myeloperoxidase deficiency, primary hypoparathyroidism, melanoma, polyoses type CDG 1, hereditary emphysema, congenital hyperthyroidism, osteogenesis imperfecta, hereditary hypofibrinogenemia, ACT deficiency, Diabetes Insipidus (DI), metaplasia (DI), nephrogenic DI, Charcot-Marie syndrome, Marian-Marie syndrome, Marie-Mar, -padi-mei's disease, neurodegenerative diseases (e.g. alzheimer's disease, parkinson's disease, amyotrophic lateral sclerosis, progressive supranuclear palsy, pick's disease), several polyglutamine neurological disorders (e.g. huntington's disease, spinocerebellar ataxia type I, spinal and bulbar muscular atrophy, dentatorubral pallidoluysian and myotonic dystrophy) and spongiform encephalopathies (e.g. hereditary creutzfeldt-jakob disease (caused by prion protein processing defects), fabry disease and stewart's syndrome), secretory diarrhoea, polycystic kidney disease, Chronic Obstructive Pulmonary Disease (COPD), xerophthalmia and sjogren's disease.

In accordance with the methods of the present invention, the compounds and compositions can be administered in any amount and by any route of administration effective to treat or reduce the severity of one or more of the following diseases: cystic fibrosis, hereditary emphysema, hereditary hemochromatosis, coagulation-fibrinolysis deficiency (e.g., protein C deficiency), hereditary angioedema type 1, lipid processing deficiency (e.g., familial hypercholesterolemia), chylomicronemia type 1, β -lipoproteinemia, lysosomal storage diseases (e.g., I-cell disease/pseudo Hurler), mucopolysaccharidosis, Sandhof/Tay-Sachs, Crigler-Najjar type II, polyendocrinosis/hyperinsulinemia, diabetes, Laron dwar, myeloperoxidase deficiency, primary hypoparathyroidism, melanoma, polyoses type CDG 1, hereditary emphysema, congenital hyperthyroidism, osteogenesis imperfecta, hereditary hypofibrinogenemia, ACT deficiency, Diabetes Insipidus (DI), metaplasia (DI), nephrogenic DI, Charcot-Marie syndrome, Marian-Marie syndrome, Marie-Mar, -padi-mei's disease, neurodegenerative diseases (e.g. alzheimer's disease, parkinson's disease, amyotrophic lateral sclerosis, progressive supranuclear palsy, pick's disease), several polyglutamine neurological disorders (e.g. huntington's disease, spinocerebellar ataxia type I, spinal and bulbar muscular atrophy, dentatorubral pallidoluysian and myotonic dystrophy) and spongiform encephalopathies (e.g. hereditary creutzfeldt-jakob disease (caused by prion protein processing defects), fabry disease and stewart's syndrome), secretory diarrhoea, polycystic kidney disease, Chronic Obstructive Pulmonary Disease (COPD), xerophthalmia and sjogren's disease.

The exact amount required will vary from subject to subject, depending on the species, age and general condition of the subject, the severity of the infection, the particular drug, the manner in which it is administered, and the like. The compounds of the present invention are preferably formulated in dosage unit form for ease of administration and uniformity of dosage. The expression "dosage unit form" as used herein denotes physically discrete pharmaceutical units, as appropriate for the patient to be treated. It will be understood, however, that the total daily amount of the compounds and compositions of the present invention will be decided by the attending physician within the scope of sound medical judgment. The specific effective dosage level for any particular patient or organism will depend upon a variety of factors including the disorder being treated and the severity of the disorder; the activity of the particular compound employed; the specific composition employed; the age, weight, general health, sex, and diet of the patient; the time of administration, the route of administration, and the rate of excretion of the particular compound employed; the duration of the treatment; drugs used in combination or concomitantly with the specific compound employed; and other factors well known in the medical arts. The term "patient" as used herein means an animal, preferably a mammal, most preferably a human.

The pharmaceutically acceptable compositions of the present invention may be administered orally, rectally, parenterally, intracisternally, intravaginally, intraperitoneally, topically (as powders, ointments or drops), buccally, as an oral or nasal spray, etc., to humans and other animals, depending on the severity of the infection being treated. In certain embodiments, the compounds of the present invention may be administered orally or parenterally at a dosage level of from about 0.01mg/kg to about 50mg/kg, preferably from about 1mg/kg to about 25mg/kg, of the subject's body weight per day, one or more times a day, to achieve the desired therapeutic effect.

Liquid dosage forms for oral administration include, but are not limited to, pharmaceutically acceptable emulsions, microemulsions, solutions, suspensions, syrups and elixirs. In addition to the active compounds, the liquid dosage forms may contain inert diluents commonly used in the art such as, for example, water or other solvents, solubilizing agents and emulsifiers such as ethyl alcohol, isopropyl alcohol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1, 3-butylene glycol, dimethylformamide, oils (in particular, cottonseed, groundnut, corn, germ, olive, castor, and sesame oils), glycerol, tetrahydrofurfuryl alcohol, polyethylene glycols and fatty acid esters of sorbitan, and mixtures thereof. Besides inert diluents, the oral compositions can also include adjuvants such as wetting agents, emulsifying and suspending agents, sweetening, flavoring, and perfuming agents.

Injectable preparations, for example sterile injectable aqueous or oleaginous suspensions, may be formulated according to the known art using suitable dispersing or wetting agents and suspending agents. The sterile injectable preparation may also be a sterile injectable solution, suspension or emulsion in a non-toxic parenterally-acceptable diluent or solvent, for example as a solution in 1, 3-butanediol. Among the acceptable carriers and solvents that may be employed are water, ringer's solution, U.S. p. and isotonic sodium chloride solution. In addition, sterile fixed oils are conventionally employed as a solvent or suspending medium. For this purpose, any bland fixed oil may be employed including synthetic mono-or di-glycerides. In addition, fatty acids, such as oleic acid, may be used in the preparation of injectables.

The injectable formulations can be sterilized, for example, by filtration through a bacterial-retaining filter, or by incorporating sterilizing agents in the form of sterile solid compositions which can be dissolved or dispersed in sterile water or other sterile injectable medium prior to use.

In order to prolong the effect of the compounds of the invention, it is often desirable to delay absorption of the compounds following subcutaneous or intramuscular injection. This can be achieved by using a liquid suspension of crystalline or amorphous material which is poorly water soluble. The rate of absorption of a compound depends on its rate of dissolution, which in turn may depend on crystal size and crystal form. Alternatively, delayed absorption of the parenterally administered compound form is achieved by dissolving or suspending the compound in an oil vehicle. Injectable depot forms are prepared by forming a microencapsulated matrix of the compound in a biodegradable polymer, such as polylactide-polyglycolide. Depending on the ratio of compound to polymer and the nature of the particular polymer employed, the release rate of the compound can be controlled. Examples of other biodegradable polymers include poly (orthoesters) and poly (anhydrides). Depot injectable formulations can also be prepared by entrapping the compound in liposomes or microemulsions which are compatible with body tissues.

Rectal or vaginal compositions are preferably suppositories which can be prepared by mixing the compounds of the invention with suitable non-irritating excipients or carriers such as cocoa butter, polyethylene glycol or a suppository wax which are solid at ambient temperature but liquid at body temperature and therefore melt in the rectum or vaginal cavity to release the active compound.

Solid dosage forms for oral administration include capsules, tablets, pills, powders and granules. In such solid dosage forms, the active compound is mixed with at least one inert, pharmaceutically acceptable excipient or carrier, for example sodium citrate or dicalcium phosphate, and/or a) fillers or extenders, for example starches, lactose, sucrose, glucose, mannitol, and silicic acid, b) binders, for example carboxymethylcellulose, alginates, gelatin, polyvinylpyrrolidone, sucrose, and acacia, c) wetting agents, for example glycerol, d) disintegrants, for example agar-agar, calcium carbonate, potato or tapioca starch, alginic acid, certain silicates, and sodium carbonate, e) dissolution retarders, for example paraffin, f) absorption accelerators, for example quaternary ammonium compounds, g) wetting agents, for example cetyl alcohol and glycerol monostearate, h) absorbents, for example kaolin and bentonite, and i) lubricants, for example talc, calcium stearate, sodium silicate, and the like, Magnesium stearate, solid polyethylene glycol, sodium lauryl sulfate and mixtures thereof. In the case of capsules, tablets and pills, the dosage forms may also comprise buffering agents.

Solid compositions of a similar type may also be employed as fillers in soft or hard-filled gelatin capsules using such excipients as lactose or milk sugar as well as polymeric polyethylene glycols and the like. Solid dosage forms such as tablets, dragees, capsules, pills and granules can be provided with coatings and shells such as enteric coatings and other coatings well known in the pharmaceutical formulating art. They may optionally contain opacifying agents and may also be of a composition that they release the active ingredient(s) only, or preferentially, in a certain portion of the intestinal tract, optionally in a delayed manner. Examples of embedding compositions that may be used include polymeric substances and waxes. Solid compositions of a similar type may also be employed as fillers in soft and hard-filled gelatin capsules using such excipients as lactose or milk sugar as well as polymeric polyethylene glycols and the like.

The active compound may also be in microencapsulated form, containing one or more of the above-mentioned excipients. Solid dosage forms such as tablets, dragees, capsules, pills and granules can be provided with coatings and shells such as enteric coatings, release controlling coatings and other coatings well known in the pharmaceutical formulating art. In such solid dosage forms, the active compound may be mixed with at least one inert diluent, for example sucrose, lactose or starch. Such dosage forms may also contain, under normal circumstances, other substances in addition to inert diluents, such as tableting lubricants and other tableting aids, for example magnesium stearate and microcrystalline cellulose. In the case of capsules, tablets and pills, the dosage forms may also comprise buffering agents. They may optionally contain opacifying agents and may also be of a composition that they release the active ingredient(s) only, or preferentially, in a certain portion of the intestinal tract, optionally in a delayed manner. Examples of embedding compositions that may be used include polymeric substances and waxes.

Dosage forms for topical or transdermal administration of the compounds of the present invention include ointments, pastes, creams, lotions, gels, powders, solutions, sprays, inhalants or patches. The active ingredient is mixed under sterile conditions with a pharmaceutically acceptable carrier and any necessary preservatives or buffers, as appropriate. Ophthalmic formulations, ear drops and eye drops are also encompassed within the scope of the present invention. In addition, the present invention encompasses the use of transdermal patches, which have the added advantage of controlling the delivery of compounds to the body. Such dosage forms may be prepared by dissolving or dispersing the compound in the appropriate medium. Absorption enhancers may also be used to increase the flux of the compound across the skin. The rate can be controlled by providing a rate controlling membrane or dispersing the compound in a polymer matrix or gel.

As generally described above, the compounds of the present invention are useful as modulators of ABC transporters. Thus, without wishing to be bound by any particular theory, the compounds and compositions are particularly useful for treating or lessening the severity of a disease, disorder or condition in which hyperactivity or inactivity of an ABC transporter has been implicated. When hyperactivity or inactivity of an ABC transporter has been implicated in a particular disease, disorder or condition, the disease, disorder or condition may also be referred to as an "ABC transporter-mediated disease, disorder or condition". Thus, in another aspect, the present invention provides methods of treating or lessening the severity of a disease, disorder or condition in which hyperactivity or inactivity of the ABC transporter has been implicated in the disease state.

The activity of compounds useful as ABC transporter modulators in the present invention can be determined according to methods generally described in the art and in the examples herein.

It will also be appreciated that the compounds and pharmaceutically acceptable compositions of the present invention may be used in combination therapy, that is, the compounds and pharmaceutically acceptable compositions may be administered simultaneously, prior to, or subsequent to one or more other desired therapeutic agents or pharmaceutical procedures. The particular combination of therapies (therapeutic agents or procedures) used in the combination regimen will take into account the compatibility of the desired therapeutic agent and/or procedure with the desired therapeutic effect to be achieved. It will also be appreciated that the therapies used may achieve the desired effect on the same condition (e.g., the compounds of the invention may be administered simultaneously with another drug used to treat the same condition), or they may achieve different effects (e.g., control of any side effects). As used herein, other therapeutic agents that are normally administered to treat or prevent a particular disease or condition are referred to as "appropriate for the disease or condition being treated.

The amount of the other therapeutic agent in the composition of the present invention will not exceed the amount normally administered in a composition containing the therapeutic agent as the only active ingredient. Preferably, the amount of the other therapeutic agent in the presently disclosed compositions will be about 50% to 100% of the content in typical compositions containing the drug as the sole therapeutically active ingredient.

The compounds of the present invention or pharmaceutically acceptable compositions thereof may also be incorporated into compositions for coating implantable medical devices, such as prostheses, prosthetic valves, vascular grafts, stents and catheters. Thus, the present invention in another aspect comprises a composition for coating an implantable device comprising a compound of the present invention as generally described above and as described in classes and subclasses herein, and a carrier suitable for coating said implantable device. In another aspect, the present invention includes an implantable device coated with a composition comprising a compound of the present invention as generally described above and in classes and subclasses herein, and a carrier suitable for coating the implantable device. Suitable coatings and general methods of making coated implantable devices are described in U.S. Pat. nos. 6,099,562, 5,886,026, and 5,304,121. The coating is typically a biocompatible polymeric material such as hydrogel polymers, polymethyldisiloxane, polycaprolactone, polyethylene glycol, polylactic acid, ethylene vinyl acetate copolymers and mixtures thereof. The coating may optionally be further covered with a surface layer of a suitable fluorosilicone, polysaccharide, polyethylene glycol, phospholipid, or combinations thereof to impart controlled release characteristics to the composition.

Another aspect of the invention relates to modulating ABC transporter activity in a biological sample or patient (e.g., in vitro or in vivo), the method comprising administering to the patient or contacting the biological sample with a compound of formula I or a composition comprising the compound. The term "biological sample" as used herein includes, without limitation, cell cultures and extracts thereof; biopsy material obtained from a mammal or an extract thereof; and blood, saliva, urine, feces, semen, tears, or other body fluids or extracts thereof.

Modulation of ABC transporter activity in a biological sample can be used for a variety of purposes known to those skilled in the art. Examples of such purposes include, but are not limited to, ABC transporter studies in biological and pathological phenomena; and comparative evaluation of novel ABC transporter modulators.

In another embodiment, there is provided a method of modulating the activity of an anion channel in vitro or in vivo comprising the step of contacting the channel with a compound of formula (I). In a preferred embodiment, the anion channel is a chloride channel or a bicarbonate channel. In other preferred embodiments, the anion channel is a chloride channel.

According to an alternative embodiment, the present invention provides a method of increasing the number of functional ABC transporters in a cell membrane, comprising the step of contacting said cell with a compound of formula (I). The term "functional ABC transporter" as used herein means an ABC transporter capable of exerting transport activity. In a preferred embodiment, the functional ABC transporter is CFTR.

According to another preferred embodiment, the activity of the ABC transporter is measured by measuring the transmembrane potential. The means for measuring the transmembrane potential in a biological sample may employ any known method known in the art, such as optical membrane potentiometry or other electrophysiological methods.

Optical Membrane potential assays employing voltage-sensitive FRET sensors as described by Gonzalez and Tsien ((R))See also，Gonzalez，J.E.and R.Y.Tsien(1995)″Voltagesensing by fluorescence resonance energy transfer in singlecells″Biophys J 69(4)：1272-80，and Gonzalez，J.E.and R.Y.Tsien(1997)″Improved indicators of cell membrane potentialthat use fluorescence resonance energy transfer″Chem Biol4(4): 269-77) in combination with an instrument for measuring changes in fluorescence, e.g. voltage/ion probe reader (VIPR) ((VIPR)See alsoConzalez, J.E., K.Oads, et al (1999) "Cell-based assays and instrumentation for screening-channel targetsDrug Discov Today 4(9)：431-439)。

These voltage-sensitive assays are based on the membrane-soluble, voltage-sensitive stain DiSBAC₂(3) Change in Fluorescence Resonance Energy Transfer (FRET) with the fluorescent phospholipid CC2-DMPE, which is attached to the outer leaflet of the plasma membrane and serves as a FRET donor. Membrane potential (V)_m) Resulting in a negatively charged DiSBAC₂(3) Redistributed across the plasma membrane and the energy transferred from the CC2-DMPE changed accordingly. The change in fluorescence emission can be made using VIPR^TMII monitoring, which is an integrated liquid processor and fluorescence detector, is designed for cell type screening in 96-or 384-well microtiter plates.

In another aspect, the present invention provides a kit for measuring the activity of an ABC transporter or a fragment thereof in a biological sample in vitro or in vivo, comprising: (i) a composition comprising a compound of formula (I) or any of the above embodiments; and (ii) guidance regarding: a) contacting the composition with a biological sample; and b) measuring the activity of the ABC transporter or the fragment thereof. In one embodiment, the kit further comprises instructions for: a) contacting the other composition with the biological sample; b) measuring the activity of the ABC transporter or a fragment thereof in the presence of the other compound; and c) comparing the ABC transporter activity in the presence of the other compound with the ABC transporter density in the presence of the composition of formula (I). In a preferred embodiment, the kit is used to measure the density of CFTR.

In order that the invention described herein may be more fully understood, the following examples are provided. It should be understood that these examples are for illustration only and are not to be construed as limiting the invention in any way.

Preparation and examples

Example 1: 2- (dimethylamino) -6- (2-methoxyphenyl) pyrimidine-4-carboxamide

Step a: 4- (2-methoxyphenyl) -2, 4-dioxobutyric acid

To a stirred solution of diethyl oxalate (16g, 110mmol) and sodium ethoxide (100mL, 21 wt.% in ethanol, 300mmol) was added dropwise a solution of 2-methoxyacetophenone (15g, 100mmol) in ethanol (100 mL). The reaction mixture was stirred at ambient temperature overnight. The resulting solution was concentrated on a rotary evaporator to approximately 50mL, followed by partitioning between diethyl ether (100mL) and water (250 mL). The aqueous layer was adjusted to pH 3 with concentrated HCl to give a suspension with a fine white precipitate, heated at 100 ℃ for 5-10 min, followed by cooling in an ice bath for 1 h. The precipitate was filtered, washed with water (20mL), and dried under high vacuum overnight to give 4- (2-methoxyphenyl) -2, 4-dioxobutyric acid.¹H NMR(300MHz，CDCl₃) δ 7.96(q, J ═ 9.0, 3.0Hz, 1H), 7.53(t, J ═ 6.0Hz, 1H), 7.47(s, 0.9H ketone type), 7.24(s, 1.1H enol type), 7.06(t, J ═ 6.0Hz, 1H), 7.00(d, J ═ 9.0Hz, 1H), 3.97(s, 3H), ESI-MS M/z 220.9(M-H)^-。

Step b: 6- (2-methoxyphenyl) -2- (methylthio) pyrimidine-4-carboxylic acid

2, 4-dioxo-4- (2-methoxy)A mixture of phenylphenyl) butyric acid (15.6g, 70mmol), 2-methyl-isothiourea sulfate (10.4g, 73.5mmol) and sodium ethoxide (26mL, 21 wt.% ethanol solution, 70mmol) in ethanol (200mL) was heated to reflux overnight. The resulting solution was reduced to approximately 50mL using a rotary evaporator and water (200mL) was added to give a suspension which was adjusted to pH > 9 with 10% NaOH. The mixture was extracted with diethyl ether (100 mL). The aqueous layer was adjusted to pH 3 with concentrated HCl solution to give a suspension, which was heated to 90 ℃ for 10 minutes. After cooling in an ice bath for 1h, a precipitate formed, filtered, washed with water (20mL) and dried overnight under vacuum to give 6- (2-methoxyphenyl) -2- (methylthio) pyrimidine-4-carboxylic acid.¹H NMR(300MHz，CDCl₃) δ 8.45(s, 1H), 8.10(dd, J ═ 7.5, 1.8Hz, 1H), 7.48(m, 1H), 7.09(t, J ═ 6.9Hz, 1H), 7.02(d, J ═ 8.7Hz, 1H), 3.94(s, 3H), 2.67(s, 3H). HPLC retention time 2.02min, 10-100% CH₃CN, 5min gradient; ESI-MS M/z 277.1(M + H)⁺.

Step c: 6- (2-methoxyphenyl) -2- (methylthio) pyrimidine-4-carboxamide

To a stirred suspension of 6- (2-methoxyphenyl) -2- (methylthio) pyrimidine-4-carboxylic acid (9.8g, 40mmol), triethylamine (33mL, 240mmol) and ammonium chloride (11g, 200mmol) in DMF (40mL) was added PyBOP (21.9g, 42mmol) in five portions. The reaction was complete within 2h according to LCMS analysis. The reaction mixture was poured into water (1.5L) and stirred for 10 minutes. The precipitate was collected by filtration and dried under high vacuum overnight to give 6- (2-methoxyphenyl) -2- (methylthio) pyrimidine-4-amide.¹H NMR (300MHz, CDCl3) Δ 8.40(s, 0.7H), 8.05(m, 1.3H), 7.74(bs, 1H), 7.44(m, 1H), 7.05(m, 2H), 5.78(bs, 1H), 3.92(s, 3H); 2.65(s, 3H). HPLC retention time 2.71min, 10-100% CH₃CN, 5min gradient; ESI-MS M/z 245.9(M + H)⁺。

Step d: 6- (2-methoxyphenyl) -2- (methylsulfinyl) pyrimidine-4-amide and 6- (2-methoxyphenyl) -2- (methylsulfonyl) pyrimidine-4-amide

To a flask containing 6- (2-methoxyphenyl) -2- (methylthio) pyrimidine-4-amide (2.75g, 8.5mmol) and 5mL of DCM was added mCPBA (2.3g, 10.2mmol, 77 wt.%, remainder 3-chlorobenzoic acid and water). After stirring at room temperature for 3h, a precipitate formed and was filtered to give 6- (2-methoxyphenyl) -2- (methylsulfinyl) pyrimidine-4-amide and 6- (2-methoxyphenyl) -2- (methylsulfonyl) pyrimidine-4-amide as a white solid (mixture of sulfones and sulfoxides). HPLC retention time 1.97min, 10-100% CH₃CN, 5min gradient; ESI-MSm/z 308.1(M + H)⁺. The crude product was used in the next step without further purification.

When an excess of oxidant mCPBA was used, the sulfone was obtained as the major product.

Step e: 2- (dimethylamino) -6- (2-methoxyphenyl) pyrimidine-4-amide (192)

6- (2-methoxyphenyl) -2- (methylsulfinyl) pyrimidine-4-amide (46mg, 0.15mmol), 1, 4-dioxane (2mL), DIPEA (80. mu.L, 0.45mmol), and dimethylamine hydrochloride (26mg, 0.3mmol) were added to a vial (8 mL). The vial was sealed and heated at 85 ℃ for two days. After the reaction solution was cooled to room temperature, the solvent was removed under vacuum in a Savant SpeedVac. The resulting residue was purified by preparative HPLC.¹H NMR(300MHz，CDCl₃)δ7.95(q，J＝7.8，1.8Hz，1H)，7.88(s，1H)，7.77(bs，1H)，7.40(m，1H)，7.04(t，J＝9.0Hz，1H)，6.98(d，J＝6.0Hz，1H)，5.78(bs，1H)，3.92(s，3H)，3.26(s6H). HPLC retention time 2.76min, 10-100% CH₃CN, 5min gradient; ESI-MS M/z 273.1(M + H)⁺。

Example 2: 2- (N-methylphenylethylamino) -6-phenylpyrimidine-4-amide.

Step a: 2, 4-dioxo-4-phenylbutyric acid

To a stirred solution of diethyl oxalate (21.9g, 150mmol) and sodium ethoxide (168mL, 21 wt.% ethanol solution, 450mmol) was added dropwise a solution of acetophenone (18g, 150mmol) in ethanol (120 mL). The reaction mixture was stirred at ambient temperature for 15 h. The reaction mixture was concentrated to approximately 70mL and partitioned between diethyl ether (50mL) and water (200 mL). The phases were separated and the aqueous layer was adjusted to pH 1 with concentrated HCl and then extracted with diethyl ether (3 × 100 mL). The combined ether extracts were MgSO₄Drying and evaporating to obtain the 2, 4-dioxo-4-phenylbutyric acid.¹H NMR(DMSO-d₆300MHz) δ 8.05(d, J ═ 7.2Hz, 1.8H enol type), 7.96(d, J ═ 6.9Hz, 0.2H ketone type), 7.69(t, J ═ 7.5Hz, 1H), 7.56(t, J ═ 8.1Hz, 2H), 7.08(s, 0.9H enol type), 4.56(s, 0.1H ketone type).¹³C NMR(DMSO-d₆，75MHz)δ190.7，170.7，163.6，135.1，134.6，129.7，128.4，98.5。

Step b: 2-methylsulfanyl-6-phenylpyrimidine-4-carboxylic acid.

A mixture of 2, 4-dioxo-4-phenylbutyric acid (19.2g, 100mmol), 2-methylisothiourea sulfate (27.8g, 100mmol) and sodium ethoxide (37mL, 21% wt. solution, 100mmol) in ethanol (200mL) was heated to reflux overnight. Removing the volatileAfter the hair is developed, water (200mL) is added to give a suspension, which is adjusted to pH > 9 with 10% NaOH. The mixture was extracted with diethyl ether (2X 100 mL). The aqueous layer was adjusted to pH 2 with concentrated HCl and extracted with diethyl ether (4 × 100 mL). The combined ether extracts were evaporated and the residue dried in vacuo to give 22.0g of 2-methylsulfanyl-6-phenylpyrimidine-4-carboxylic acid as a yellow solid.¹H NMR(DMSO-d₆，300MHz)δ8.24(dd，J＝7.8，1.8Hz，2H)，8.12(s，1H)，7.58(m，3H)，2.64(s，3H)。¹³C NMR(DMSO-d₆75MHz) delta 172.8, 165.7, 165.6, 157.9, 135.8, 132.5, 129.8, 128.0, 14.6. HPLC retention time 1.95min, 10-100% CH₃CN, 5min running. ESI-MS M/z 247.1(M + H)⁺。

Step c: 2- (methylthio) -6-phenylpyrimidine-4-carboxamide

N-methylmorpholine (2.31mL, 21mmol) was added dropwise with a syringe to a mixture of 2-methylsulfanyl-6-phenylpyrimidine-4-carboxylic acid (3.45g, 14mmol), ammonium chloride (4.27g, 80mmol), HOBt (2.84g, 21mmol) and EDC (4.0g, 21mmol) in DMF (40mL) at room temperature. The reaction solution was stirred at room temperature for 3h, then DMF was removed by rotary evaporation. Subjecting to column chromatography (SiO)₂DCM/EtOAc 5: 1) to give 3.23g of 2- (methylthio) -6-phenylpyrimidine-4-amide as a white solid.¹H NMR(DMSO-d₆300MHz) delta 8.25(m, 2H), 8.13(s, 1H), 7.59(m, 3H), 2.62(s, 3H). HPLC retention time 2.71min, 10-100% CH₃CN, 5min gradient. ESI-MS M/z 246.1(M + H)⁺。

Step d: 2- (methylsulfonyl) -6-phenylpyrimidine-4-carboxamide

To a flask containing 2-methylsulfanyl-6-phenylpyrimidine-4-amide (2.0g, 8.2mmol) and 100mL of DCM was added mCPBA (30mmol, 5.18g, 77 wt.%, remainder 3-chlorobenzoic acid and water). After stirring at room temperature for 3h, a white precipitate formed, filtered off and washed on the filter with cold DCM (20mL) to give 2- (methanesulfonyl) -6-phenylpyrimidine-4-amide as a white solid.¹H NMR(DMSO-d₆300MHz) δ 8.67(s, 1H), 8.50(s, 1H), 8.34(dd, J ═ 7.8, 1.5Hz, 2H), 8.18(s, 1H), 7.63(m, 3H), 3.60(s, 3H). HPLC retention time 2.10min, 10-100% CH₃CN, 5min running. ESI-MS M/z 278.1(M + H)⁺。

Step e: 2- (N-Methylphenylethylamino) -6-phenylpyrimidine-4-amide (title compound).

2-methanesulfonyl-6-phenylpyrimidine-4-amide (70mg, 0.25mmol), 1, 4-dioxane (4mL), DIPEA (0.3mL, 2.0mmol), and N-methylphenethylamine (82mg, 0.6mmol) were added to an 8mL vial. The vial was sealed and heated at 85 ℃ for two days. The reaction mixture was allowed to cool to room temperature and then the solvent was removed by centrifugal evaporation. The residue was purified by preparative HPLC to give 43mg of 2- (N-methylphenylethylamino) -6-phenylpyrimidine-4-amide.¹H NMR(CDCl₃300MHz) δ 8.14(m, 2H), 7.74(s, 1H), 7.67(br s, 1H), 7.47(m, 3H), 7.25(m, 5H), 5.79(br s, 1H), 3.95(t, J ═ 6.9Hz, 2H), 3.21(s, 3H), 2.97(t, J ═ 6.9Hz, 2H). HPLC retention time 3.25min, 10-100% CH₃CN, 5min running. ESI-MS M/z 333.3(M + H)⁺。

Example 3: 2-diethylamino-6- (2, 6-dimethoxy-phenyl) -pyrimidine-4-carboxylic acid amide

Step a: 2-methylsulfanyl-6-oxo-1, 6-dihydro-pyrimidine-4-carboxylic acid

Neat diethyl oxalacetate (23.5g, 125mmol) was added to a solution of sodium hydroxide (15g, 0.375mol, 3eq) in water (150mL) to give a yellow solution. To the stirred solution was added solid S-methylisothiouronium sulfate (17.4g, 62.5mmol, 0.5 eq). The mixture was stirred for 15 hours. The pH was adjusted to 1 by the addition of concentrated HCl (40mL, 0.48mol) and the resulting light orange-pink suspension was stirred vigorously for 2 hours and then filtered. The filter cake was washed with water (100mL) and dried under high vacuum to give 2-methylsulfanyl-6-oxo-1, 6-dihydro-pyrimidine-4-carboxylic acid as an off-white solid.¹H NMR(500MHz，DMSO-d₆)δ13.22(br s，1H)，6.60(s，1H)，3.37(br s，1H)，2.53(s，3H)；ESI-MS m/z 186.9(M+H)⁺。

Step b: 6-chloro-2-methylsulfanyl-pyrimidine-4-carboxylic acid methyl ester

A250 mL flask was charged with 2-methylsulfanyl-6-oxo-1, 6-dihydro-pyrimidine-4-carboxylic acid (11.6g, 62.5mmol), toluene (60mL), thionyl chloride (60mL, 0.8mol, 13eq.), DMF (0.3mL) and pyridine (1mL), and the resulting suspension was stirred and heated to reflux. The suspension cleared after 30 min. After refluxing for 3 hours, the mixture was concentrated in vacuo and co-evaporated with additional toluene (30 mL). The biphasic brown residue was dissolved in chloroform (100mL), cooled to 0 ℃, and DIEA (56mL, 0.31mol, 5eq.) was added dropwise over 20min, followed by methanol (12.7mL, 0.31mol, 5eq.) over 5min. The mixture was warmed to room temperature and poured into saturated aqueous sodium bicarbonate (350 mL). The organic layer was washed with saturated aqueous ammonium chloride (350mL) and dried (Na)₂SO₄) Filtering, in vacuumConcentrating to brown semi-solid. Drying under high vacuum overnight afforded 6-chloro-2-methylsulfanyl-pyrimidine-4-carboxylic acid methyl ester, which was used in the next step without further purification.¹H NMR(500MHz，CDCl₃)δ7.60(s，1H)，3.98(s，3H)，2.60(s，3H)；ESI-MS m/z 218.9(M+H)⁺。

Step c: 6-chloro-2-methylsulfanyl-pyrimidine-4-carboxylic acid amides

A500 mL flask was charged with 6-chloro-2-methylsulfanyl-pyrimidine-4-carboxylic acid methyl ester (4.37g, 20mmol) and chloroform (30 mL). Ammonia (57mL, 7M methanol solution, 0.4mol, 20eq.) was added while stirring. HPLC analysis after 20 minutes showed no starting methyl ester remaining. The mixture was sparged with nitrogen for 30min, then concentrated in vacuo and dried under high vacuum to give 6-chloro-2-methylsulfanyl-pyrimidine-4-carboxylic acid amide (4.21g, quantitative) in sufficient purity for the next step.¹H NMR(500MHz，CDCl₃)δ7.76(s，1H)，7.60(brs，1H)，5.97(br s，1H)，2.61(s，3H)；ESI-MS m/z 204.1(M+H)⁺。

Step d: 6- (2, 6-dimethoxy-phenyl) -2-methylsulfanyl-pyrimidine-4-carboxylic acid amide

A7 mL vial with a Teflon screw cap was charged with 2, 6-dimethoxyphenyl-boronic acid (218mg, 1.2mmol, 2eq.) and dichloro-bis- (diphenylphosphinoferrocene) -palladium-dichloromethane adduct (49mg, 60. mu. mol, 10 mol%). Under nitrogen atmosphere (O)₂In a glove box at 0.5%) a vial was charged with 6-chloro-2-methylsulfanyl-pyrimidine-4-carboxylic acid amide (122mg, 0.6mmol) in degassed DMFDME (1/1, 4.5mL) and degassed 1M aqueous potassium phosphate (1.0mL, 1mmol, 1.7 eq.). The vial was sealed, removed from the glove box, and agitated in a 75 ℃ constant temperature shaker for 16 hours. The mixture was cooled and then diluted with chloroform (15mL) and washed with brine (20mL, 2X 10mL) and 1M aqueous potassium carbonate (10 mL). The organic layer (6- (2, 6-dimethoxy-phenyl) -2-methylsulfanyl-pyrimidine-4-carboxylic acid amide in chloroform) was then used directly in the next step.

Step e: 6- (2, 6-dimethoxy-phenyl) -2-methanesulfonyl-pyrimidine-4-carboxylic acid amide

To a 50mL Falcon centrifuge tube containing a solution of 6- (2, 6-dimethoxy-phenyl) -2-methylsulfanyl-pyrimidine-4-carboxylic acid amide (assuming 0.6mmol) from the previous step in chloroform (15mL) was charged solid mCPBA (0.57g, 70% purity, the remaining 30% consisting of water and benzoic acid, 2.4mmol, 4eq., ca.25 wt% water) and the mixture was shaken at room temperature overnight. To the resulting light brown-orange suspension was added dimethyl sulfide (0.22mL, 3mmol, 5eq.) to quench the residual MCPBA, and the mixture was shaken for 2 hours. The mixture was then washed with 1M aqueous sodium hydroxide (10mL), brine (10mL) and dried (Na)₂SO₄) Filtered and concentrated to dryness in a centrifugal evaporator. The residue, 6- (2, 6-dimethoxy-phenyl) -2-methanesulfonyl-pyrimidine-4-carboxylic acid amide, was then dissolved in DMSO (1.8mL) and used directly for the next reaction.

Step f: 2-diethylamino-6- (2, 6-dimethoxy-phenyl) -pyrimidine-4-carboxylic acid amide

A4 mL screw cap vial was charged with 6- (2, 6-dimethoxy-phenyl)) -2-methanesulfonyl-pyrimidine-4-carboxylic acid amide (assuming 0.3mmol) in DMSO (0.9 mL). A DMSO solution of diethylamine and N-methyl-morpholine (NMM) (0.3mL, 3M each, 0.9mmol, 3eq.) was added, the vial sealed and heated at 75 ℃ for 22 hours. The mixture was then subjected to preparative HPLC to give 18mg (18%) of 2-diethylamino-6- (2, 6-dimethoxy-phenyl) -pyrimidine-4-carboxylic acid amide.¹H NMR(300MHz，CDCl₃) δ 7.79(br s, 1H), 7.30(t, J ═ 8.3Hz, 1H), 7.26(s, 1H), 6.61(d, J ═ 8.3Hz, 2H), 6.16(br s, 1H), 3.74(s, 6H), 3.66(q, J ═ 6.9Hz, 4H), 1.22(t, J ═ 6.9Hz, 6H); HPLC retention time 2.85min., 10-100% CH₃CN, 5min gradient; ESI-MS M/z 330.2(M + H)⁺。

Example 4: other Compounds

The remaining compounds described in table 1 were produced using the procedures described herein and known synthetic methods.

Table 2 includes physical data distinguishing each of the synthesized compounds.

Compound number	LC-MSM+1	LC-RTmin	NMR
				1	324.	3.39
2	308.1	2.81
				3	390.	3.48
4	286.1	2.45
				5	314.2	3.2	1H NMR(300MHz，CDCl3)δ7.98-7.93(m，2H)，7.73(br s，1H)，7.37(dt，J＝1.7，7.8Hz，1H)，7.03(t，J＝7.5Hz，1H)，6.97(d，J＝8.3Hz，1H)，5.66(br s，1H)，4.13(q，J＝6.9Hz，2H)，3.70(q，J＝7.0Hz，4H)，1.48(t，J＝6.9Hz，3H)，1.24(t，J＝7.0Hz，6H)
6	318.1	3.14
				7	256.1	2.93	1H NMR(300MHz，CDCl3)δ8.15(m，2H)，7.73(m，2H)，7.46(m，3H)，5.68(s，1H)，3.80(m，2H)，3.26(s，3H)，1.25(t，J＝9.0Hz，3H).
8	342.2	3.02	1H NMR(300MHz，CDCl3)δ7.87(s，1H)，7.71(brs，1H)，7.52(d，J＝2.7Hz，1H)，6.98-6.87(m，3H)，5.74(br s，1H)，3.89-3.81(m，10H)，1.76-1.57(m，6H)
				9	243.1	3.08
10	338.1	3.38
				11	310.2	3.44	1H NMR(300MHz，CDCl3)δ7.69(br s，1H)，7.34(s，1H)，7.28-7.11(m，3H)，5.54(br s，1H)，3.84(app t，J＝5.3Hz，4H)，2.33(s，3H)，2.30(s，3H)，1.73-1.51(m，6H)
12	334.1	3.24
				13	309.	2.92
14	268.	2.99
				15	310.1	3.01
16	282.1	3.18
				17	368.2	3.36
18	319.	2.46
				19	314.2	2.71
20	314.2	3.21
				21	290.1	3.39	1H NMR(300MHz，CDCl3)δ7.69(br s，1H)，7.57(d，J＝3.7Hz，1H)，7.49(s，1H)，6.78(dd，J＝1.0，3.7Hz，1H)，5.48(br s，1H)，3.65(q，J＝7.0

Compound number	LC-MSM+1	LC-RTmin	NMR
							Hz，4H)，2.53(s，3H)，1.24(t，J＝7.0Hz，6H)
22	313.2	2.99
				23	299.3	3.89
24	362.2	3.32	1H NMR(300MHz，CDCl3)δ7.94(dd，J＝1.4，7.7Hz，1H)，7.81(d，J＝0.5Hz，1H)，7.68(br s，1H)，7.43-7.15(m，4H)，7.13-6.81(m，1H)，5.72(br s，1H)，3.63(q，J＝7.0Hz，4H)，1.18(t，J＝7.0Hz，6H)
				25	272.1	2.89
26	269.1	3.32
				27	268.	2.81
28	420.	3.33
				29	354.1	3.32
30	256.9	3.03
				31	272.	2.83
32	296.2	3.37
				33	300.	2.9
34	284.2	3.26	1H NMR(300MHz，CDCl3)δ8.13(m，2H)，7.72(s，2H)，7.46(m，3H)，5.60(s，1H)，3.70(m，2H)，3.63(m，2H)，1.72(m，4H)，1.26(t，J＝7.2Hz，3H)，0.98(t，J＝7.2Hz，3H).
				35	328.2	3.04
36	310.2	3.45
				37	372.	3.59
38	332.	3.25	1H-NMR(CDCl3，300MHz)δ8.14(m，2H)，7.74(s，1H)，7.67(br s，1H)，7.47(m，3H)，7.23(m，5H)，5.79(br s，1H)，3.95(t，J＝6.9Hz，2H)，3.21(s，3H)，2.97(t，J＝6.9Hz，2H).
				39	340.2	3.36
40	310.2	3.38
				41	282.2	2.91
42	408.	2.99
				43	312.	2.46
44	340.	2.73
				45	400.	3.18
46	280.	3.14

Compound number	LC-MSM+1	LC-RTmin	NMR
				47	285.1	3.65
48	284.1	2.58	1H NMR(300MHz，CDCl3)δ7.79(s，1H)，7.65(m，3H)，7.37(m，1H)，7.02(m，1H)，5.97(m，2H)，5.46(s，1H)，5.28(d，J＝15.6Hz，1H)，5.16(d，J＝8.7Hz，1H)，4.16(d，J＝5.7Hz，2H)，3.87(s，3H).
				49	390.	2.96
50	332.	3.26
				51	271.1	3.23
52	360.2	3.47
				53	356.	3.41
54	330.	3.27
				55	312.2	3.47
56	284.2	3.32
				57	266.3	2.71
58	265.9	1.99
				59	354.	3.05
60	404.	3.13
				61	284.2	3.25
62	327.2	2.47
				63	371.	3.11
64	256.9	3.03
				65	342.2	3.07
66	290.1	3.21
				67	288.1	3.27
68	296.	3.38
				69	294.	3.15
70	302.1	3.22
				71	271.3	3.15
72	350.1	3.12	1H NMR(300MHz，CDCl3)δ7.77(d，J＝7.5Hz，1H)，7.67(br s，1H)，7.64-7.47(m，3H)，7.35(s，1H)，5.60(br s，1H)，3.96-3.77(m，4H)，1.73-1.52(m，6H)
				73	324.	3.63
74	300.2	3.08
				75	338.1	3.27
76	342.2	3.4	1H NMR(300MHz，CDCl3)δ7.88-7.73(m，3H)，

Compound number	LC-MSM+1	LC-RTmin	NMR
							7.29-7.11(m，1H)，6.92(d，J＝8.4Hz，1H)，5.87(br s，1H)，3.87(s，3H)，3.70(q，J＝6.9Hz，4H)，2.92(heptet，J＝6.8Hz，1H)，1.25(app t，J＝6.9Hz，12H)
77	272.1	2.69
				78	326.	3.32
79	266.	2.91
				80	328.1	2.45
81	326.1	3.4
				82	330.2	2.85	1H NMR(300MHz，CDCl3)δ7.79-7.72(m，3H)，7.66(s，1H)，6.95(d，J＝8.1Hz，1H)，5.71(br s，1H)，3.97(s，3H)，3.94(8，3H)，3.72(q，J＝7.0Hz，4H)，1.27(t，J＝7.0Hz，6H)
83	267.	2.29
				84	354.1	3.32
85	356.2	3.31
				86	310.	3.43
87	310.	3.42	1H-NMR(CDCl3，300MHz)δ8.11(m，2H)，7.72(s，1H)，7.71(br s，1H)，7.45(m，3H)，5.77(br s，1H)，3.69(t，J＝7.2Hz，2H)，3.60(d，J＝6.3Hz，2H)，1.71(m，2H)，1.20(m，1H)，0.97(t，J＝7.2Hz，3H)，0.54(m，2H)，0.33(m，2H).
				88	316.1	2.44	1H NMR(300MHz，CDCl3)δ7.76(m，1H)，7.65(m，2H)，7.36(m，J＝8.1Hz 1H)，7.00(d，J＝7.5Hz1H)，6.04(s，1H)，5.62(s，1H)，3.87(s，3H)，3.62(m，2H)，3.53(d，J＝6.0Hz，2H)，3.36(s，3H)，1.96(m，2H).
89	340.2	3.31
				90	332.	2.01
91	298.9	3.92
				92	394.	3.34	1H-NMR(CDCl3，300MHz)δ8.11(m，2H)，7.83(s，1H)，7.49(br s，1H)，7.43(m，3H)，7.29(m，10H)，5.58(br s，1H)，5.04(br s，2H)，4.89(br s，2H)
93	318.	3.27
				94	288.1	3.39
95	350.1	3.3

Compound number	LC-MSM+1	LC-RTmin	NMR
				96	442.	3.11	1H-NMR(CDCl3，300MHz)δ8.13(m，2H)，7.78(s，1H)，7.76(s，1H)，7.67(m，2H)，7.47(m，5H)，5.85(br s，1H)，4.91(d，J＝12.6Hz，2H)，3.51(m，2H)，2.14(m，2H)，2.07(br s，1H)，1.89(d，J＝12.6Hz，2H).
97	360.9	4.2
				98	304.1	3.3
99	354.	3.01
				100	300.2	2.53
101	348.	2.73	1H-NMR(CDCl3，300MHz)δ8.12(m，2H)，7.79(s，1H)，7.64(br s，1H)，7.48(m，3H)，7.30(m，5H)，5.85(br s，1H)，5.16(t，J＝5.7Hz，1H)，3.98(d，J＝5.7Hz，2H)，3.17(s，3H).
				102	272.	2.61
103	242.1	2.47	1H-NMR(CDCl3，300MHz)δ8.09(m，2H)，7.91(brs 1H)，7.80(s，1H)，7.46(m，3H)，5.23(br s，1H)，3.09(d，J＝5.1Hz，3H)，3.03(d，J＝5.1Hz，3H).
				104	355.	4.05	1H NMR(300MHz，CDCl3)δ8.14(m，1H)，7.72(m，2H)，7.43(m，3H)，5.59(s，1H)，3.63(m，4H)，1.57(m，6H)，0.99(d，J＝6.6Hz，12H).
105	330.1	3.06
				106	350.1	3.4
107	306.1	3.11
				108	330.2	2.77	1H NMR(300MHz，CDCl3)δ7.79(br s，1H)，7.30(t，J＝8.3Hz，1H)，7.26(s，1H)，6.61(d，J＝8.3Hz，2H)，6.16(br s，1H)，3.74(s，6H)，3.66(q，J＝6.9Hz，4H)，1.22(t，J＝6.9Hz，6H)
109	300.	3.17
				110	300.	3.12
111	307.1	2.97
				112	282.9	3.3
113	310.	3.44
				114	296.2	3.33
115	346.1	3.3
				116	298.2	3.55
117	298.9	3.51

Compound number	LC-MSM+1	LC-RTmin	NMR
				118	312.	3.55
119	271.1	3.48	H NMR(400MHz，DMSO-d6)δ8.16(m，2H)，8.06(s，1H)，7.80(s，1H)，7.58(s，1H)，7.55(m，3H)，3.75(q，J＝6.9Hz，4H)，1.19(t，J＝6.5Hz，6H)
				120	298.2	3.37
121	298.2	3.4
				122	300.1	3.34
123	316.1	1.53
				124	342.2	3.02
125	306.1	2.95
				126	305.3	3.23
127	326.2	3.22
				128	322.	2.75
129	310.	3.09
				130	307.1	2.98
131	302.2	3.44
				132	405.1	3.28
133	318.1	3.09
				134	312.	2.86
135	310.	3.07
				136	296.9	3.43
137	330.1	3.14
				138	272.1	2.5
139	414.	2.6
				140	350.1	3.57
141	327.1	3.9	H NMR(400MHz，DMSO-d6)δ8.17(m，2H)，7.53(m，3H)，7.18(s，1H)，3.67(s，4H)，3.45(m，2H)，3.28(q，J＝7.0Hz，2H)，1.16(m，12H)
				142	312.	3.2
143	388.	2.93	1H-NMR(CDCl3，300MHz)δ8.10(m，2H)，7.72(s，1H)，7.67(br d，J＝3.6Hz，1H)，7.46(m，3H)，7.27(m，5H)，5.83(br d，J＝3.6Hz，1H)，5.65(d，J＝13.2Hz，2H)，3.41(m，2H)，2.80(s，2H)，1.10(m，4H).
				144	338.1	3.34
145	282.1	2.42

Compound number	LC-MSM+1	LC-RTmin	NMR
				146	374.2	3.36
147	304.1	3.31
				148	336.	3.67
149	296.2	3.27
				150	338.	3.47
151	306.1	3.53	1H NMR(300MHz，CDCl3)δ7.70-7.60(m，2H)，7.63(s，1H)，6.91(app t，J＝8.3Hz，1H)，5.57(br s，1H)，3.72(q，J＝7.0Hz，4H)，1.26(t，J＝7.0Hz，6H)
				152	366.1	3.23
153	294.1	3.2
				154	252.	2.5	1H-NMR(CDCl3，300MHz)δ8.11(m，2H)，7.88(s，1H)，7.71(br s，1H)，7.47(m，3H)，5.83(br s，1H)，5.47(br s，1H)，4.32(m，2H)，2.24(t，J＝2.7Hz，1H).
155	362.1	2.7
				156	300.2	2.55
157	338.7	3.94
				158	300.2	3.16	1H NMR(300MHz，CDCl3)δ8.03(s，1H)，7.99(dt，J＝6.5，2.1Hz，1H)，7.02(s，1H)，4.76(s，2H)，4.67(heptet，J＝6.9Hz，1H)，3.69(q，J＝7.0Hz，4H)，3.43(q，J＝7.4Hz，2H)，1.31t，J＝6.1Hz，3H)，1.26-1.14(m，6H)，1.22(d，J＝6.9Hz，6H)
159	284.2	3.23
				160	288.1	3.27
161	326.2	3.25
				162	308.1	2.94
163	306.1	3.13
				164	319.3	3.57
165	296.2	3.28
				166	324.2	3.52
167	284.1	2.54
				168	312.2	3.1
169	286.	2.85
				170	338.1	3.42
171	327.3	4.24

Compound number	LC-MSM+1	LC-RTmin	NMR
				172	330.2	3.01
173	324.1	2.61
				174	332.2	2.74
175	314.2	3.48
				176	324.	2.6
177	381.	2.69
				178	284.	2.61
179	229.1	2.58
				180	383.	3.15
181	326.	2.48	1H-NMR(CDCl3，300MHz)δ8.11(m，2H)，7.71(s，1H)，7.69(br s，1H)，7.45(m，3H)，5.82(br s，1H)，4.89(d，J＝13.2Hz，2H)，3.75(t，J＝6.6Hz，2H)，2.96(m，2H)，1.84(d，J＝12.0Hz，2H)，1.75(m，1H)，1.57(q，J＝6.3Hz，2H)，1.27(m，2H).
				182	366.1	3.37
183	254.9	2.98
				184	326.2	3.63
185	350.1	3.32	1H NMR(300MHz，CDCl3)δ8.37(br s，1H)，8.30(d，J＝7.6Hz，1H)，7.77-7.64(m，3H)，7.59(t，J＝7.9Hz，1H)，5.62(br s，1H)，3.95-3.85(m，4H)，1.77-1.60(m，6H)
				186	298.2	3.41
187	433.3	3.57
				188	296.	3.25
189	298.1	2.76
				190	358.2	3.01
191	244.1	1.98
				192	300.1	3.44
193	328.	2.48
				194	326.2	3.26
195	454.1	3.37
				196	326.	3.62
197	340.	3.38
				198	326.2	3.26
199	350.1	3.65
				200	338.1	3.51

Compound number	LC-MSM+1	LC-RTmin	NMR
				201	312.2	3.59
202	350.1	3.36
				203	298.1	2.74
204	266.	2.72
				205	312.	3.17
206	298.	3.41
				207	298.2	3.18
208	272.	2.77
				209	314.	2.71
210	338.1	3.23
				211	314.1	2.51
212	314.2	3.21
				213	366.1	3.35
214	326.1	3.3
				215	243.1	2.81	1H-NMR(CDCl3，300MHz)δ8.09(m，2H)，7.78(s，1H)，7.71(br s，1H)，7.47(m，3H)，5.72(br s，1H)，5.25(br s，1H)，3.55(m，2H)，1.30(t，J＝7.5Hz，3H).
216	294.	2.67
				217	330.2	2.98	1H NMR(300MHz，CDCl3)δ7.89(s，1H)，7.76(brs，1H)，7.56(d，J＝2.8Hz，1H)，6.98-6.89(m，3H)，5.83(br s，1H)，3.85(s，3H)，3.82(s，3H)，3.69(q，J＝7.0Hz，4)，1.24(t，J＝7.0Hz，6H)
218	347.	2.82
				219	318.1	3.57
220	284.	2.68
				221	296.2	3.47
222	310.	3.28
				223	328.2	2.97
224	354.2	3.4
				225	338.1	3.29
226	338.1	3.57
				227	326.1	3.04
228	316.	2.45
				229	301.2	2.93
230	284.2	3.43

Compound number	LC-MSM+1	LC-RTmin	NMR
				231	258.1	2.31
232	296.2	3.33
				233	338.1	3.31

Determination of

Assays for detecting and measuring Δ F508-CFTR modulating properties of compounds

A. Membrane potential optical method for determining regulatory property of compound delta F508-CFTR

Optical Membrane potential assays employing voltage-sensitive FRET sensors as described by Gonzalez and Tsien ((R))See also，Gonzalez，J.E.and R.Y.Tsien(1995)″Voltagesensing by fluorescence resonance energy transfef in singlecells″Biophys J 69(4)：1272-80，and Gonzalez，J.E.and R.Y.Tsien(1997)″Improved indicators of cell membrane potentialthat use fluorescence resonance energy transfer″Chem Biol4(4): 269-77) in combination with an instrument for measuring changes in fluorescence, e.g. voltage/ion probe reader (VIPR) ((VIPR)See alsoGonzalez, J.E., K.Oads, et al (1999) "Cell-based assays and instrumentation for screening-channel targetsDrug Discov Today 4(9)：431-439)。

These voltage-sensitive assays are based on the membrane-soluble, voltage-sensitive stain DiSBAC₂(3) Change in Fluorescence Resonance Energy Transfer (FRET) with the fluorescent phospholipid CC2-DMPE, which is attached to the outer leaflet of the plasma membrane and serves as a FRET donor. Membrane potential (V)_m) Resulting in a negatively charged DiSBAC₂(3) Redistributed across the plasma membrane and the energy transferred from the CC2-DMPE changed accordingly. The change in fluorescence emission can be made using VIPR^TMII monitoring, which is an integrated liquid processor and fluorescence detector, is designed for 96-or 384-well microtiter plate based screening.

1. Identification of modulating Compounds

To identify small molecules that modulate transport defects associated with Δ F508-CFTR, a single-addition HTS assay format was developed. Cells were incubated in serum-free medium at 37 ℃ for 16 hours in the presence or absence (negative control) of test compound. As a positive control, cells plated in 384-well plates were incubated at 27 ℃ for 16 hours to "temperature-adjust" Δ F508-CFTR. The cells were then washed 3 times with a solution of Krebs Ringer (Krebs Ringer) and loaded with a voltage-sensitive stain. To activate Δ F508-CFTR, 10 μ M forskolin and CFTR enhancer genistein (20 μ M) and no Cl were added to each well^-And (4) a culture medium. Cl-free^-Addition of media promotes Cl in response to activation of Δ F508-CFTR^-Efflux, the resulting membrane depolarization was monitored optically using FRET-based voltage-sensing agents.

2. Identification of enhancer compounds

To identify enhancers for Δ F508-CFTR, a dual-addition HTS assay format was developed. During the first addition, each well was added with Cl-free with or without test compound^-And (4) a culture medium. After 22 seconds, Cl-free containing 2-10. mu.M forskolin was performed^-Second addition of media to activate af 508-CFTR. Extracellular Cl after two additions^-Concentration 28mM, which promotes Cl in response to activation of Δ F508-CFTR^-Efflux, the resulting membrane depolarization was monitored optically using FRET-based voltage-sensing agents.

3. Solutions of

Bath solutionLiquid #1 (mM): NaCl 160, KCl 4.5, CaCl₂ 2，MgCl₂ 1，HEPES10，pH 7.4(NaOH)。

Chlorine-free bath solution: the chloride salt in bath solution #1 was replaced with gluconate.

CC 2-DMPE: stock solutions of 10mM were made in DMSO and stored at-20 ℃.

DiSBAC₂(3): stock solutions of 10mM were made in DMSO and stored at-20 ℃.

4. Cell culture

Optical measurements of membrane potential were performed using NIH3T3 mouse fibroblasts stably expressing AF 508-CFTR. At 175cm²Cells were maintained at 37 ℃ in 5% CO in culture flasks₂And 90% humidity, and Dulbecco's modified Eagle's Medium supplemented with 2mM glutamine, 10% fetal bovine serum, 1X NEAA, beta-ME, 1X penicillin/streptomycin, and 25mM HEPES. For all optical assays, cells were seeded at 30,000/well in 384-well matrigel-coated plates, incubated at 37 ℃ for 2 hours, and then at 27 ℃ for 24 hours for reinforcer assays. For the conditioning assay, cells were incubated with and without compound at 27 ℃ or 37 ℃ for 16-24 hours.

B. Electrophysiological assay for determining the Δ F508-CFTR modulating properties of a compound

1. Using cell assay methods

Ussing laboratory experiments were performed on polarized epithelial cells expressing Δ F508-CFTR to further characterize modulators of Δ F508-CFTR identified in the optical assay. FRT to be grown on CostarSnapwell cell culture insert^ΔF508-CFTREpithelial cells were mounted in a using chamber (physiological Instruments, inc., San Diego, CA) and monolayers were continuously shorted using a voltage clamp system (Department of biotechnology, University of Iowa, IA, and, physiological Instruments, inc., San Diego, CA). Applying 2mV pulsesThe transepithelial electrical resistance is measured. Under these conditions, FRT epithelium exhibits 4K Ω/cm²Or a resistance of the above. The solution was maintained at 27 ℃ and air was blown in. Electrode offset potential and fluid resistance were adjusted using a batteryless insert. Under these conditions, the current reflects Cl in the top film^-Flow through Δ F508-CFTR. Digital I acquisition Using MP100A-CE interface and AcqKnowledge software (v3.2.6; BIOPAC Systems, Santa Barbara, Calif.)_SC。

2. Identification of modulating Compounds

A typical protocol employs a substrate lateral to apical membrane Cl^-A concentration gradient. To establish this gradient, normal ringer's solution was used for the basolateral membrane, while the apical NaCl was replaced by equimolar sodium gluconate (titrated to pH7.4 with NaOH), yielding large Cl spans across the epithelium^-A concentration gradient. All experiments were performed with the entire monolayer. To fully activate Δ F508-CFTR, forskolin (10 μ M) and the PDE inhibitor IBMX (100 μ M) were applied followed by the addition of the CFTR enhancer genistein (50 μ M).

As observed in other cell types, incubation of FRT cells stably expressing af 508-CFTR at low temperatures increases the functional density of CFTR in the plasma membrane. To determine the activity of the modulatory compounds, cells were incubated with 10 μ M of test compound at 37 ℃ for 24 hours, followed by 3 washes and then recorded. cAMP-and Genistein-mediated Compound-treated cells I_SCNormalized to the 27 ℃ and 37 ℃ controls, expressed as a percentage of activity. Pre-incubation of cells with modulating compounds significantly increased cAMP-and genistein-mediated I compared to 37 ℃ controls_SC。

3. Identification of enhancer compounds

A typical protocol employs a substrate lateral to apical membrane Cl^-A concentration gradient. To establish this gradient, the basolateral membrane was permeabilized with nystatin (360. mu.g/ml) using a normal ringer's solution, while the apical NaCl was equimolar to gluconic acidSodium substitution (titration with NaOH to pH7.4) gave large Cl spanning the epithelium^-A concentration gradient. All experiments were performed 30 minutes after nystatin permeabilization. Forskolin (10 μ M) and all test compounds were added to both sides of the cell culture insert. The efficacy of the putative AF 508-CFTR enhancer was compared to that of the known enhancer, genistein.

4. Solutions of

Substrate outer solution (mM): NaCl (135), CaCl₂(1.2)，MgCl₂(1.2)，K₂HPO₄(2.4)，KHPO₄(0.6), N-2-hydroxyethylpiperazine-N' -2-ethanesulfonic acid (HEPES) (10), glucose (10). The solution was titrated with NaOH to pH 7.4.

Apical solution (mM): like the solution on the outside of the substrate, NaCl was replaced with sodium gluconate (135).

5. Cell culture

Fisher rat epithelial (FRT) cells expressing Δ F508-CFTR (FRT) were used^ΔF508-CFTR) Ussing laboratory experiments were performed on putative Δ F508-CFTR modulators identified from our optical assays. Cells were cultured on Costar Snapwell cell culture insert at 37 ℃ and 5% CO₂In Coon's modified Ham's F-12 medium supplemented with 5% fetal bovine serum, 100U/ml penicillin and 100. mu.g/ml streptomycin for 5 days. Cells were incubated at 27 ℃ for 16-48 hours to adjust Δ F508-CFTR before use to characterize the potentiating properties of the compounds. To determine the activity of the modulatory compounds, cells were incubated with and without the compound for 24 hours at 27 ℃ or 37 ℃.

6. Whole cell recording

Macroscopic AF 508-CFTR current (I) was monitored in NIH3T3 cells stably expressing AF 508-CFTR, conditioned by temperature and test compound, using punch-patch whole cell recordings_ΔF508). Briefly, an Axomatch 200B patch-clamp amplifier (Axon instruments Inc., Foster City, Calif.) was used at room temperatureIs as follows_ΔF508Voltage clamp record of (1). All recordings were taken at a sampling frequency of 10kHz and low pass filtered at 1 kHz. The pipette has a resistance of 5-6M Ω when filled with the intracellular solution. Under these recording conditions, the Cl at room temperature was calculated^-Reversal potential (E)_C1) Was-28 mV. All records had a seal resistance > 20G Ω and a series resistance < 15M Ω. Pulsing, data acquisition and analysis were performed using a PC equipped with a Digidata 1320A/D interface and Clampex 8 (Axoinstruments Inc.). The bath contained < 250 μ L saline and was continuously perfused using a gravity driven perfusion system at a rate of 2 ml/min.

7. Identification of modulating Compounds

To determine the activity of the modulatory compounds to increase the density of functional af 508-CFTR in plasma membranes, we measured the current density 24 hours after the modulatory compounds were treated using the punch-patch-recording technique described above. To fully activate Δ F508-CFTR, 10 μ M forskolin and 20 μ M genistein were added to the cells. Under our recording conditions, the current density after 24 hours of incubation at 27 ℃ was higher than that observed after 24 hours of incubation at 37 ℃. These results are consistent with the known effect of low temperature incubation on Δ F508-CFTR density in plasma membranes. To determine the effect of a modulating compound on CFTR current density, cells were incubated with 10 μ M of test compound at 37 ℃ for 24 hours and the current density (% activity) was compared to 27 ℃ and 37 ℃ controls. Prior to recording, cells were washed 3 times with extracellular recording medium to remove any remaining test compound. Preincubation with 10 μ M of modulating compound significantly increased cAMP-and genistein-dependent current compared to 37 ℃ control.

8. Identification of enhancing Compounds

The research on increasing macroscopic delta F508-CFTR Cl in NIH3T3 cells stably expressing delta F508-CFTR by using a perforating-patch-recording technology is also carried out^-Electric current (I)_ΔF508) The ability of the cell to perform. Fortifier elicitation from optical assay_ΔF508Dose-dependent increase in (A), efficacy and workThe effect is similar to that of optical assay. In all cells examined, the reversal potential before and during the application of the reinforcer was around-30 mV, which is the calculated E_C1(-28mV)。

9. Solutions of

Intracellular solution (mM): cs-aspartate (90), CsCl (50), MgCl₂(1) HEPES (10) and 240. mu.g/ml amphotericin-B (pH adjusted to 7.35 with CsOH).

Extracellular solution (mM): N-methyl-D-glucamine (NMDG) -Cl (150), MgCl₂(2)，CaCl₂(2) HEPES (10) (pH adjusted to 7.35 with HCl).

10. Cell culture

Whole cell recordings were performed using NIH3T3 mouse fibroblasts stably expressing AF 508-CFTR. At 175cm²Cells were maintained at 37 ℃ in 5% CO in culture flasks₂And 90% humidity, and Dulbecco's modified Eagle's Medium supplemented with 2mM glutamine, 10% fetal bovine serum, 1X NEAA, beta-ME, 1X penicillin/streptomycin, and 25mM HEPES. For whole cell recording, 2,500-5,000 cells were seeded on poly-L-lysine coated glass coverslips, incubated at 27 ℃ for 24-48 hours, and then used to test for enhancer activity; incubation at 37 ℃ with or without the modulating compound was used to measure the activity of the modulator.

11. Single channel recording

The single channel activity of temperature-adjusted Δ F508-CFTR stably expressed in NIH3T3 cells and the activity of enhancer compounds were observed using excised membrane sheets with inside-out turning. Briefly, single channel active voltage clamp recordings were performed at room temperature using an Axomatch 200B patch-clamp amplifier (Axon instruments Inc.). All recordings were taken at a sampling frequency of 10kHz and low pass filtered at 400 Hz. The patch pipette was made of Corning KovarSealing #7052 glass (World Precision Instruments, Inc., Sarasota, FL) and filled with cellsThe outer solution has a resistance of 5-8M omega. After excision 1mM MG-ATP and 75nM cAMP-dependent protein kinase catalytic subunit (PKA; Promega Corp. Madison, Wis.) were added to activate Δ F508-CFTR. After the channel is stable in motion, the gravity is used for driving the micro-perfusion system to perfuse the membrane. The influent was placed near the membrane, resulting in complete solution exchange within 1-2 seconds. To maintain Δ F508-CFTR activity during rapid perfusion, a non-specific phosphatase inhibitor F was added to the bath solution^-(10mM NaF). Under these recording conditions, the channel activity remained constant throughout the patch recording period (up to 60 minutes). The current generated by the movement of positive charges from the intracellular solution to the extracellular solution (the anions moving in the opposite direction) is shown as positive current. Pipette potential (V)_p) The concentration is maintained at 80 mV.

Channel activity was analyzed from patches containing less than or equal to 2 active channels. The maximum number of simultaneous openings during the course of the experiment determines the number of active channels. To determine single channel current amplitude, data recorded from 120 sec Δ F508-CFTR activity was filtered "off-line" at 100Hz and then used to construct a full-point amplitude histogram that was fitted to a multi-Gaussian function using Bio-Patch analysis software (Bio-Logic Comp. France). Total microscopic Current and open probability (P) were determined from 120 second channel Activity₀)。P₀Is made by using Bio-batch software or slave P₀I/I (N), where I is the average current, I is the single-channel current amplitude, and N is the number of active channels in the diaphragm.

12. Solutions of

Extracellular solution (mM): NMDG (150), aspartic acid (150), CaCl₂(5)，MgCl₂(2) And HEPES (10) (adjusted pH to 7.35 with Tris base).

Intracellular solution (mM): NMDG-Cl (150), MgCl₂(2) EGTA (5), TES (10) and Tris base (14) (pH adjusted to 7.35 with HCl).

13. Cell culture

Use of NIH3T3 mice stably expressing AF 508-CFTR for fibroblast growthThe fibroblasts were subjected to excision membrane patch clamp recordings. At 175cm²Cells were maintained at 37 ℃ in 5% CO in culture flasks₂And 90% humidity, and Dulbecco's modified Eagle Medium supplemented with 2mM glutamine, 10% fetal bovine serum, 1X NEAA, beta-ME, 1X penicillin/streptomycin, and 25mM HEPES. For single channel recording, 2,500-5,000 cells were seeded on poly-L-lysine coated glass coverslips and incubated at 27 ℃ for 24-48 hours before use.

Table 3 contains the results of the measurements of the compounds of table 1. The ranges of potency determined in table 3 are as follows: the potency of +++ corresponds to EC50 of less than 1.0. mu.M, + EC50 of between 1.0. mu.M and 5.0. mu.M, + EC50 of greater than 5.0. mu.M. The range of efficacy determined in table 3 is as follows: , + ++ corresponds to a power greater than 100, + corresponds to a power between 100 and 50, + corresponds to a power less than 50.

TABLE 3

Compound No.	EC50	Efficacy of
			217	+++	++
218	++	+++
			219	++	+
220	+	+
			221	++	+
222	+	++
			223	++	+++
224	+	+
			225	+++	+++
226	++	++
			227	+	++
228	+	++
			229	++	++
230	++	+++
			231	+	++
232	++	+
			233	+++	++

Other embodiments

It is to be understood that while the invention has been described in conjunction with the detailed description thereof, the foregoing description is intended to illustrate and not limit the scope of the invention, which is defined by the scope of the appended claims. Other aspects, advantages, and modifications are within the scope of the following claims.

Claims (51)

1. A compound of the formula:

wherein

Each Ra is independently H, an optionally substituted aliphatic, an optionally substituted aryl, an optionally substituted aralkyl, an optionally substituted heteroaralkyl, an optionally substituted heteroaryl, an optionally substituted cycloaliphatic, or an optionally substituted cycloheteroaliphatic;

each Rb is independently an optionally substituted aliphatic, an optionally substituted aryl, an optionally substituted aralkyl, an optionally substituted heteroaralkyl, an optionally substituted heteroaryl, an optionally substituted cycloaliphatic, an optionally substituted cycloheteroaliphatic, an optionally substituted heteroaryl, a,Wherein w is 1, 2, 3, 4 or 5, the phenyl group is optionally substituted with 1-4 Re, or Ra and Rb together with the nitrogen atom to which they are bound form an optionally substituted heterocycloaliphatic, optionally substituted heteroaryl;

each Rc is independently H, an optionally substituted heterocycloaliphatic, an optionally substituted cycloaliphatic, or an unsubstituted aliphatic;

each Rd is independently H, an optionally substituted aliphatic, an optionally substituted aryl, an optionally substituted aralkyl, an optionally substituted heteroaralkyl, an optionally substituted heteroaryl, an optionally substituted cycloaliphatic, an optionally substituted cycloheteroaliphatic, or Rc and Rd taken together with the nitrogen atom to which they are bound form an optionally substituted heterocycloaliphatic;

ring a is aryl or heteroaryl, each optionally substituted with 1-4 Re;

each Re is independently carboxy, amino, nitro, alkylcarbonylamino, cycloalkylcarbonylamino, (cycloalkyl) alkylcarbonylamino, arylcarbonylamino, aralkylcarbonylamino, (heterocycloalkyl) carbonylamino, (heterocycloalkyl) alkylcarbonylamino, heteroarylcarbonylamino, heteroaralkylcarbonylamino, sulfonyl, sulfinyl, sulfanyl, sulfoxy, sulfamoyl, sulfonylamino, ketal, carbamoyl, cyano, halo, urea, thiourea, haloalkyl or-Z-Rf, wherein each alkyl, cycloalkyl, heterocycloalkyl, aryl and heteroaryl portion of the Re substituent is optionally substituted with 1-3 Rg, or

Two Re on adjacent a ring atoms form, together with the a ring atom to which they are bonded, a heterocycloaliphatic ring;

each Z is absent, -O-or-S-;

each Rf is independently hydrogen, alkyl, carboxyalkyl, hydroxyalkyl, alkenyl, alkynyl, cycloalkyl, (cycloalkyl) alkyl, heterocycloalkyl, (heterocycloalkyl) alkyl, aryl, heteroaryl, aralkyl, heteroaralkyl, aroyl, heteroaroyl, alkoxycarbonyl, alkylcarbonyl, aminocarbonyl, or acyl, wherein each alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl, and heteroaryl moiety on the Rf substituent is optionally substituted with 1-3 Rg;

each Rg is independently halogen, cyano, alkoxy, hydroxy, nitro, haloalkyl, and alkyl;

provided that it is

When Rd is alkyl substituted by 4-methyl-1-piperazinyl, then Ring A is not 3, 4, 5-trimethoxyphenyl;

ring a is not 7-chloro-2- (4-fluorophenyl) pyrazolo [1, 5-a ] pyridin-3-yl or 7-cyclopentylamino-2- (4-fluorophenyl) pyrazolo [1, 5-a ] pyridin-3-yl;

when Rc is H, Ra and Rb are both alkyl, and ring a is oxazolyl, then Rd is not {1- [ (3, 5-difluorophenyl) methyl) ] -2-hydroxy-4- (1H-pyrazol-3-yl) butyl); and is

When ring a is phenyl, Ra and Rb together with the nitrogen to which they are bound form pyrrolidine, and Rd is substituted alkyl, then Rc is not H.

2. The compound of claim 1, wherein ring a is aryl optionally substituted with 1-4 Re.

3. The compound of claim 2, wherein said aryl is optionally substituted with 1-4 alkyl, hydroxyalkyl, haloalkyl, alkoxy, halogen, hydroxy, alkoxycarbonyl, aryloxy, S (O)₂Alkyl, cyano, alkylcarbonylamino, methylenedioxy or acyl.

4. The compound of claim 3, wherein said aryl is phenyl.

5. The compound of claim 1, wherein ring a is heteroaryl.

6. The compound of claim 5, wherein said heteroaryl is optionally substituted with 1-4 alkyl, hydroxyalkyl, haloalkyl, alkoxy, halogen, hydroxy, alkoxycarbonyl, aryloxy, sulfoxy, or acyl groups.

7. The compound of claim 6, wherein ring a is thiophene, pyrimidinyl, benzothiophene, or pyridyl, furanyl.

8. The compound of claim 1, wherein Ra is H.

9. The compound of claim 1, wherein Ra is an optionally substituted aliphatic.

10. The compound of claim 9, wherein Ra is optionally substituted alkyl.

11. The compound of claim 10, wherein Ra is hydroxyalkyl, alkoxyalkyl, (heterocycloalkyl) alkyl, (cycloalkyl) alkyl, or alkoxycarbonylalkyl.

12. The compound of claim 9, wherein Ra is an optionally substituted alkenyl or an optionally substituted alkynyl.

13. The compound of claim 1, wherein Ra is optionally substituted aralkyl or optionally substituted heteroaralkyl.

14. The compound of claim 1, wherein Rb is an optionally substituted aliphatic.

15. The compound of claim 14, wherein Rb is an optionally substituted alkyl.

16. The compound of claim 15, wherein Rb is hydroxyalkyl, alkoxyalkyl, (heterocycloalkyl) alkyl, (cycloalkyl) alkyl, or alkoxycarbonylalkyl.

17. The compound of claim 1, wherein Rb is an optionally substituted alkenyl or an optionally substituted alkynyl.

18. The compound of claim 1, wherein Rb is an optionally substituted aralkyl or an optionally substituted heteroaralkyl.

19. The compound of claim 1, wherein Ra and Rb together form an optionally substituted heterocycloaliphatic or an optionally substituted heteroaryl.

20. The compound of claim 19, wherein Ra and Rb together form an optionally substituted heterocycloaliphatic.

21. The compound of claim 20, wherein Ra and Rb form an optionally substituted heterocycloalkyl.

22. The compound of claim 21, wherein said heterocycloalkyl is optionally substituted with 1-3 of halo, haloalkyl, alkyl, alkoxycarbonyl, alkylcarbonyl, hydroxyalkyl, sulfonyl, sulfinyl, aminocarbonyl, cyano, sulfoxy, acetal, ketal, optionally substituted aryl, optionally substituted alkoxy, optionally substituted aralkyl, optionally substituted aroyl, or optionally substituted heteroaryl.

23. The compound of claim 22, wherein the heterocycloalkyl is a piperidine, piperazine, morpholino, thiomorpholino, tetrahydropyridinyl, decahydroisoquinolinyl, pyrrolidinyl, thiazolidinyl, or azetidinyl ring, wherein each ring is optionally substituted with 1-3 of halo, haloalkyl, alkyl, alkoxycarbonyl, alkylcarbonyl, hydroxyalkyl, sulfonyl, sulfinyl, aminocarbonyl, cyano, sulfoxy, acetal, ketal, an optionally substituted aryl, an optionally substituted alkoxy, an optionally substituted aralkyl, an optionally substituted aroyl, or an optionally substituted heteroaryl.

24. The compound of claim 20, wherein Ra and Rb form an optionally substituted heteroaryl.

25. The compound of claim 24, wherein said heteroaryl is optionally substituted with 1-3 of halo, haloalkyl, alkyl, alkoxycarbonyl, alkylcarbonyl, hydroxyalkyl, sulfonyl, sulfinyl, aminocarbonyl, cyano, sulfoxy, acetal, ketal, optionally substituted aryl, optionally substituted alkoxy, optionally substituted aralkyl, optionally substituted aroyl, or optionally substituted heteroaryl.

26. The compound of claim 24, wherein the heteroaryl is a tetrahydroisoquinolinyl, perhydroisoquinoline, imidazolyl, or pyrazolyl ring, wherein each ring is optionally substituted with 1-3 of halo, haloalkyl, alkyl, alkoxycarbonyl, alkylcarbonyl, hydroxyalkyl, sulfonyl, sulfinyl, aminocarbonyl, cyano, sulfoxy, acetal, ketal, an optionally substituted aryl, an optionally substituted alkoxy, an optionally substituted aralkyl, an optionally substituted aroyl, or an optionally substituted heteroaryl.

27. The compound of claim 1, wherein Rd is H or an optionally substituted aliphatic.

28. The compound of claim 27, wherein Rd is H.

29. The compound of claim 1, wherein Ra and Rb are aliphatic.

30. The compound of claim 1, wherein Ra and Rb are alkyl.

31. The compound of claim 30, wherein each Ra and Rb is independently methyl, ethyl, propyl, or butyl.

32. The compound of claim 31, wherein Ra and Rb are both methyl, ethyl, propyl, or butyl.

33. The compound of claim 32, wherein Ra and Rb are both ethyl.

34. The compound of claim 1, wherein ring a is substituted with at least one Re.

35. The compound of claim 34, wherein ring a is substituted with one Re ortho to the point of attachment between ring a and the pyrimidine.

36. The compound of claim 34, wherein ring a is substituted with one Re meta to the point of attachment between ring a and the pyrimidine.

37. The compound of claim 34, wherein ring a is substituted with one Re para to the point of attachment between ring a and the pyrimidine.

38. The compound of claim 1, wherein ring a is substituted with two Re.

39. The compound of claim 38, wherein the two Re are ortho and meta to the point of attachment between ring a and the pyrimidine.

40. The compound of claim 1, wherein Rc is H.

41. The compound of claim 1, wherein Rd is H.

42. The compound of claim 1, wherein Rc and Rd are both H.

43. A compound selected from the group consisting of:

2-azepan-1-yl-6-phenyl-pyrimidine-4-amides

2- (4-acetyl-4-phenyl-1-piperidinyl) -6-phenyl-pyrimidine-4-carboxamide

2- (cyclopropylmethylamino) -N-methyl-6-phenyl-pyrimidine-4-carboxamide

2- (4-methyl-1-piperidinyl) -6-phenyl-pyrimidine-4-carboxamides

6- (3-methoxyphenyl) -2-morpholino-pyrimidine-4-amide

2- (2-Furanylmethyl-amino) -6- (3-methoxyphenyl) -pyrimidine-4-carboxamide

2- (butyl-propyl-amino) -6-phenyl-pyrimidine-4-carboxamide

N-methyl-2-methylamino-6-phenyl-pyrimidine-4-carboxamide

2- [4- (4-chlorophenyl) -4-hydroxy-1-piperidinyl ] -6-phenyl-pyrimidine-4-carboxamide

2-ethylamino-6- (3-methoxyphenyl) -pyrimidine-4-amide

6- (3, 5-dichlorophenyl) -2- (1-piperidinyl) pyrimidine-4-carboxamides

2-diethylamino-6- (6-methoxy-3-pyridinyl) -pyrimidine-4-carboxamide

2-diisobutylamino-6-phenyl-pyrimidine-4-amides

6- (3-furyl) -2- (1-piperidinyl) pyrimidine-4-carboxamides

2- (2-Furanylmethyl-amino) -6- (4-methoxyphenyl) -pyrimidine-4-carboxamide

2- (methyl-pentyl-amino) -6-phenyl-pyrimidine-4-amide

6- (2, 3-dichlorophenyl) -2-diethylamino-pyrimidine-4-carboxamide

3- (6-carbamoyl-2-diethylamino-pyrimidin-4-yl) benzoic acid isopropyl ester

6- (2, 3-difluorophenyl) -2- (1-piperidinyl) pyrimidine-4-carboxamides

2- (2, 6-dimethylmorpholin-4-yl) -6-phenyl-pyrimidine-4-amide

2-azepan-1-yl-N, N-dimethyl-6-phenyl-pyrimidine-4-carboxamide

6-phenyl-2-pyrrolidin-1-yl-pyrimidine-4-amides

6- (2, 5-Dimethoxyphenyl) -2- (1-piperidinyl) pyrimidine-4-carboxamide

2- { 4-hydroxy-4- [3- (trifluoromethyl) phenyl ] -1-piperidinyl ] -6-phenyl-pyrimidine-4-carboxamide

6- (4-methoxyphenyl) -2- (1, 2, 3, 6-tetrahydropyridin-1-yl) pyrimidine-4-amide

6- (2, 5-dichlorophenyl) -2- (1-piperidinyl) pyrimidine-4-carboxamides

6-benzothien-3-yl-2-diethylamino-pyrimidine-4-carboxamide

2- (cyclopropylmethylamino) -6-phenyl-pyrimidine-4-carboxamide

2-diethylamino-6- (2, 6-dimethoxyphenyl) -pyrimidine-4-carboxamide

2-diethylamino-6- (3-ethoxyphenyl) -pyrimidine-4-carboxamide

2- (allyl-methyl-amino) -6-phenyl-pyrimidine-4-carboxamide

2, 5-dihydro-1H-pyrrol-1-yl- [2- (2, 5-dihydro-1H-pyrrol-1-yl) -6-phenyl-pyrimidin-4-yl ] -methanone

2- (cyclopropylmethyl-propyl-amino) -6- (3-methoxyphenyl) -pyrimidine-4-amide

2-dibenzylamino-6-phenyl-pyrimidine-4-amides

2- (butyl-ethyl-amino) -6-phenyl-pyrimidine-4-carboxamide

6- (3-fluorophenyl) -2- (1-piperidinyl) pyrimidine-4-carboxamides

2-diethylamino-6- (3, 5-difluorophenyl) -pyrimidine-4-carboxamide

6- (5-isopropyl-2-methoxy-phenyl) -2- (1-piperidinyl) pyrimidine-4-amide

2-diethylamino-N-ethyl-6-phenyl-pyrimidine-4-carboxamide

2- [ (4-carbamoyl-6-phenyl-pyrimidin-2-yl) -methyl-amino ] acetic acid ethyl ester

2- (ethyl- (2-hydroxyethyl) amino) -6- (3-methoxyphenyl) -pyrimidine-4-amide

2-diethylamino-6- (2-fluorophenyl) -pyrimidine-4-carboxamide

2- (1-piperidinyl) -6- [2- (trifluoromethyl) phenyl ] -pyrimidine-4-carboxamides

2- [4- (4-methoxyphenyl) sulfonylpiperazin-1-yl ] -6-phenyl-pyrimidine-4-amide

6- (2-methoxyphenyl) -2- (tetrahydrofuran-2-ylmethyl-amino) pyrimidine-4-amide

6- (4-ethoxyphenyl) -2- (1-piperidinyl) pyrimidine-4-carboxamides

2-cyclopentylamino-6-phenyl-pyrimidine-4-amides

2-dipropylamino-6-phenyl-pyrimidine-4-amides

(2-diethylamino-6-phenyl-pyrimidin-4-yl) - (1-piperidinyl) methanones

6- (4-methoxyphenyl) -2- (1-piperidinyl) pyrimidine-4-amide

2-dimethylamino-6-phenyl-pyrimidine-4-carboxamide

2-diethylamino-6- (3-methoxyphenyl) -pyrimidine-4-carboxamide

2-diethylamino-6- (5-methyl-2-thienyl) -pyrimidine-4-carboxamide

2-allylamino-6- (3-methoxyphenyl) -pyrimidine-4-carboxamide

2- [ [1- (3, 4-dimethoxyphenyl) cyclopentyl ] methylamino ] -6-phenyl-pyrimidine-4-amide

2-diethylamino-6- (2-methoxyphenyl) -pyrimidine-4-carboxamide

2-azepan-1-yl-N-methyl-6-phenyl-pyrimidine-4-carboxamide

2- (1-piperidinyl) -6- (p-tolyl) pyrimidine-4-carboxamides

2-diethylamino-6- (5-fluoro-2-methoxy-phenyl) -pyrimidine-4-carboxamide

6- (3-methoxyphenyl) -2- (1, 2, 3, 6-tetrahydropyridin-1-yl) pyrimidine-4-amide

2- (cyclopropylmethylamino) -6- (3-methoxyphenyl) -pyrimidine-4-carboxamide

2-diethylamino-6- (4-isobutylphenyl) -pyrimidine-4-carboxamide

2-diethylamino-6- (5-isopropyl-2-methoxy-phenyl) -pyrimidine-4-carboxamide

6-phenyl-2- (4-phenyl-1, 2, 3, 6-tetrahydropyridin-1-yl) -pyrimidine-4-amide

2-diethylamino-6- (3, 4-dimethylphenyl) -pyrimidine-4-carboxamide

2-diethylamino-6-phenyl-pyrimidine-4-carboxamides

6- (3-chlorophenyl) -2-diethylamino-pyrimidine-4-carboxamide

2-diethylamino-6- (3, 4-dimethoxyphenyl) -pyrimidine-4-carboxamide

2-diethylamino-6- [3- (trifluoromethyl) phenyl ] -pyrimidine-4-carboxamide

6- (3, 4-dichlorophenyl) -2-diethylamino-pyrimidine-4-carboxamide

6- (2-methoxyphenyl) -2- (1-piperidinyl) pyrimidine-4-amides

6-phenyl-2- (1, 2, 3, 4-tetrahydroisoquinolin-2-yl) pyrimidine-4-carboxamide

2-diethylamino-6- (m-tolyl) pyrimidine-4-carboxamide

6-phenyl-2- (1-piperidinyl) pyrimidine-4-amides

6- (5-chloro-2-methoxy-phenyl) -2- (1-piperidinyl) pyrimidine-4-amide

2-diethylamino-6- (2, 5-dimethoxyphenyl) -pyrimidine-4-carboxamide

6-phenyl-2- (4-propyl-1-piperidinyl) -pyrimidine-4-amides

6- (4-isopropylphenyl) -2- (1-piperidinyl) pyrimidine-4-amides

2-dimethylamino-6- (3-methoxyphenyl) -pyrimidine-4-carboxamide

2-diethylamino-6- (4-fluoro-3-methyl-phenyl) -pyrimidine-4-carboxamide

2-diethylamino-6- (p-tolyl) pyrimidine-4-carboxamide

N, N-dimethyl-6-phenyl-2- (1-piperidinyl) pyrimidine-4-amides

2-diethylamino-6- [3- (hydroxymethyl) phenyl ] -pyrimidine-4-carboxamide

2-diethylamino-6- (4-ethylphenyl) -pyrimidine-4-carboxamide

2- (ethyl- (2-hydroxyethyl) amino) -6- (2-methoxyphenyl) -pyrimidine-4-amide

6-phenyl-2-thiazolidin-3-yl-pyrimidine-4-amides

2- (1, 2, 3, 4, 4a, 5, 6, 7, 8, 8 a-decahydroisoquinolin-2-yl) -6-phenyl-pyrimidine-4-carboxamide

6- (2-fluoro-3-methoxy-phenyl) -2- (1-piperidinyl) pyrimidine-4-carboxamide

2- [4- (4-chlorophenyl) -1, 2, 3, 6-tetrahydropyridin-1-yl ] -6-phenyl-pyrimidine-4-amide

2-diethylamino-6- (4-methoxyphenyl) -pyrimidine-4-carboxamide

2-allylamino-6- (4-methoxyphenyl) -pyrimidine-4-amide

2- (2-furylmethyl-amino) -N-methyl-6-phenyl-pyrimidine-4-carboxamide

2-diethylamino-6- (2-phenoxyphenyl) -pyrimidine-4-carboxamide

6-benzothien-3-yl-2- (1-piperidinyl) pyrimidine-4-amides

2- (cyclopropylmethyl-propyl-amino) -6- (2-methoxyphenyl) -pyrimidine-4-amide

6-phenyl-2- (1, 4-thiazinan-4-yl) pyrimidine-4-amides

2- (1, 4-dioxa-8-azaspiro [4.5] decan-8-yl) -6-phenyl-pyrimidine-4-carboxamide

2-cyclohexylamino-6-phenyl-pyrimidine-4-amides

2- (methyl-phenethyl-amino) -6-phenyl-pyrimidine-4-amide

6-phenyl-2-propylamino-pyrimidine-4-amides

6- (2, 3-dimethylphenyl) -2- (1-piperidinyl) pyrimidine-4-amides

2- (methyl-prop-2-ynyl-amino) -6-phenyl-pyrimidine-4-carboxamide

6- (2-fluorophenyl) -2- (1-piperidinyl) pyrimidine-4-carboxamides

2- (2-furylmethyl-methyl-amino) -6-phenyl-pyrimidine-4-carboxamide

2-benzylamino-6-phenyl-pyrimidine-4-amides

6- (2, 5-dichlorophenyl) -2-diethylamino-pyrimidine-4-carboxamide

3- (6-carbamoyl-2-diethylamino-pyrimidin-4-yl) benzoic acid methyl ester

6- (3, 5-difluorophenyl) -2- (1-piperidinyl) pyrimidine-4-carboxamides

6-phenyl-2-tert-butylamino-pyrimidine-4-amides

2- (1-piperidinyl) -6- [4- (trifluoromethoxy) phenyl ] -pyrimidine-4-carboxamide

2- (benzyl-ethyl-amino) -6-phenyl-pyrimidine-4-carboxamide

6- (2, 4-dichlorophenyl) -2- (1-piperidinyl) pyrimidine-4-carboxamides

2- (2-furylmethyl-amino) -6-phenyl-pyrimidine-4-carboxamide

6- (6-methoxy-3-pyridyl) -2- (1-piperidyl) pyrimidine-4-amide

2- [ ethyl- [2- (2-pyridyl) ethyl ] amino ] -6-phenyl-pyrimidine-4-carboxamide

2- (2-hydroxyethyl-propyl-amino) -6-phenyl-pyrimidine-4-amide

2-morpholino-6-phenyl-pyrimidine-4-amides

2-diethylamino-6- (2, 4-dimethoxypyrimidin-5-yl) -pyrimidine-4-carboxamide

6- (2-ethoxyphenyl) -2- (1-piperidinyl) pyrimidine-4-carboxamides

6- (3-methoxyphenyl) -2-methylamino-pyrimidine-4-carboxamide

2- (4-cyano-4-phenyl-1-piperidinyl) -6-phenyl-pyrimidine-4-carboxamide

2- [3- (diethylcarbamoyl) -1-piperidinyl ] -6-phenyl-pyrimidine-4-carboxamides

2-ethylamino-6- (4-methoxyphenyl) -pyrimidine-4-amide

6- (3, 4-dichlorophenyl) -2- (1-piperidinyl) pyrimidine-4-carboxamides

6- (4-cyanophenyl) -2- (1-piperidinyl) pyrimidine-4-carboxamides

2- (1-piperidinyl) -6- [3- (trifluoromethyl) phenyl ] -pyrimidine-4-carboxamides

2- (ethyl- (2-methylprop-2-enyl) amino) -6-phenyl-pyrimidine-4-carboxamide

2- [ bis (2-ethoxyethyl) amino ] -6-phenyl-pyrimidine-4-carboxamide

2-diethylamino-6- {3- (trifluoromethoxy) phenyl ] -pyrimidine-4-carboxamide

4- (6-carbamoyl-2-diethylamino-pyrimidin-4-yl) benzoic acid methyl ester

6-benzothien-2-yl-2-diethylamino-pyrimidine-4-carboxamide

6- (2-phenoxyphenyl) -2- (1-piperidinyl) pyrimidine-4-carboxamides

2- (cyclopropylmethyl-propyl-amino) -6-phenyl-pyrimidine-4-carboxamide

2-azepan-1-yl-6- (4-methoxyphenyl) -pyrimidine-4-amide

2-diethylamino-6- (4-ethylsulfonylphenyl) -pyrimidine-4-carboxamide

6- (4-methyl-2-thienyl) -2- (1-piperidinyl) pyrimidine-4-carboxamides

6- (4-chlorophenyl) -2-diethylamino-pyrimidine-4-carboxamide

3- [ 6-carbamoyl-2- (1-piperidinyl) pyrimidin-4-yl ] benzoic acid isopropyl ester

2-diethylamino-6- (4-ethoxyphenyl) -pyrimidine-4-carboxamide

2-diethylamino-6- (2, 3-dimethylphenyl) -pyrimidine-4-carboxamide

6- (m-tolyl) -2- (1-piperidinyl) pyrimidine-4-carboxamides

2- (1-piperidinyl) -6- [2- (trifluoromethoxy) phenyl ] -pyrimidine-4-carboxamide

2- (cyclopropylmethyl-propyl-amino) -N, N-dimethyl-6-phenyl-pyrimidine-4-carboxamide

2- (ethyl-methyl-amino) -6-phenyl-pyrimidine-4-carboxamide

2- [4- (4-fluorobenzoyl) -1-piperidinyl ] -6-phenyl-pyrimidine-4-carboxamide

2-dibutylamino-6-phenyl-pyrimidine-4-carboxamide

6- (4-fluoro-3-methyl-phenyl) -2- (1-piperidinyl) pyrimidine-4-amide

2-isopropylamino-6-phenyl-pyrimidine-4-amides

6- (5-fluoro-2-methoxy-phenyl) -2- (1-piperidinyl) pyrimidine-4-carboxamide

6- (3-ethoxyphenyl) -2- (1-piperidinyl) pyrimidine-4-carboxamides

6- (3, 4-Dimethoxyphenyl) -2- (1-piperidinyl) pyrimidine-4-amide

2- (methyl- (3-pyridylmethyl) amino) -6-phenyl-pyrimidine-4-carboxamide

2-diethylamino-6- (4-fluorophenyl) -pyrimidine-4-amide

2-diethylamino-6- (4-methyl-2-thienyl) -pyrimidine-4-carboxamide

N-methyl-6-phenyl-2- (1-piperidinyl) pyrimidine-4-amides

2-diethylamino-6- (2-ethoxyphenyl) -pyrimidine-4-carboxamide

2-isobutylamino-6-phenyl-pyrimidine-4-amides

2-diisopentylamino-6-phenyl-pyrimidine-4-carboxamide

2- (cyanomethyl-methyl-amino) -6-phenyl-pyrimidine-4-carboxamide

2-methylamino-6-phenyl-pyrimidine-4-carboxamide

2- [4- (2-hydroxyethyl) -1-piperidinyl ] -6-phenyl-pyrimidine-4-amide

6- (3, 5-dichlorophenyl) -2-diethylamino-pyrimidine-4-carboxamide

2- (ethyl- (2-hydroxyethyl) amino) -N-methyl-6-phenyl-pyrimidine-4-amide

2- (4-benzyl-1-piperidinyl) -6-phenyl-pyrimidine-4-amides

2-diethylamino-N, N-diethyl-6-phenyl-pyrimidine-4-carboxamide

2- (benzyl-butyl-amino) -6-phenyl-pyrimidine-4-carboxamide

2- [4- (4-chlorobenzoyl) -1-piperidinyl ] -6-phenyl-pyrimidine-4-carboxamide

2-diethylamino-6- (2, 4-difluorophenyl) -pyrimidine-4-carboxamide

1- (4-carbamoyl-6-phenyl-pyrimidin-2-yl) piperidine-3-carboxylic acid ethyl ester

2- (6, 7-dimethoxy-1, 2, 3, 4-tetrahydroisoquinolin-2-yl) -6-phenyl-pyrimidine-4-carboxamide

6- (2, 3-dichlorophenyl) -2- (1-piperidinyl) pyrimidine-4-carboxamides

6- (3-methoxyphenyl) -2-prop-2-ynylamino-pyrimidine-4-amide

2- [3- (hydroxymethyl) -1-piperidinyl ] -6-phenyl-pyrimidine-4-carboxamides

2-diethylamino-6- (4-isopropylphenyl) -pyrimidine-4-carboxamide

6- (3-methoxyphenyl) -2- (3-methoxypropylamino) pyrimidine-4-amide

2-diethylamino-N-methyl-6-phenyl-pyrimidine-4-carboxamide

6- (3-cyanophenyl) -2- (1-piperidinyl) pyrimidine-4-carboxamides

6- (5-chloro-2-methoxy-phenyl) -2-diethylamino-pyrimidine-4-carboxamide

2- (4-benzyl-4-hydroxy-1-piperidinyl) -6-phenyl-pyrimidine-4-carboxamide

N-benzyl-2-diethylamino-6-phenyl-pyrimidine-4-carboxamide

2- (benzyl-methyl-amino) -6-phenyl-pyrimidine-4-amides

2-diethylamino-6- (3-fluorophenyl) -pyrimidine-4-carboxamide

1- (4-carbamoyl-6-phenyl-pyrimidin-2-yl) piperidine-4-carboxylic acid ethyl ester

2- (1H-imidazol-1-yl) -6-phenyl-pyrimidine-4-amide

2- (2, 5-dihydro-1H-pyrrol-1-yl) -6-phenyl-pyrimidine-4-carboxamide

2-azacyclooctan-1-yl-6-phenyl-pyrimidine-4-amides

6- (2, 4-Dimethoxyphenyl) -2- (1-piperidinyl) pyrimidine-4-amide

2-dimethylamino-6- (4-methoxyphenyl) -pyrimidine-4-carboxamide

2-azepan-1-yl-6- (3-methoxyphenyl) -pyrimidine-4-amide

2- [4- (2-oxo-1, 3-dihydrobenzoimidazol-1-yl) -1-piperidinyl ] -6-phenyl-pyrimidine-4-amide

2- (1-piperidinyl) -6- [3- (trifluoromethoxy) phenyl ] -pyrimidine-4-carboxamide

2-dimethylamino-6- (2-methoxyphenyl) -pyrimidine-4-carboxamide

2-allylamino-6- (2-methoxyphenyl) -pyrimidine-4-carboxamide

2- (ethyl-propyl-amino) -6-phenyl-pyrimidine-4-carboxamide

6-phenyl-2- (1H-pyrazol-1-yl) pyrimidine-4-amides

2-diallylamino-6-phenyl-pyrimidine-4-amides

2- (hexyl-methyl-amino) -6-phenyl-pyrimidine-4-carboxamide

2-diethylamino-6- (2, 4-dimethoxyphenyl) -pyrimidine-4-carboxamide

6-phenyl-2-prop-2-ynylamino-pyrimidine-4-amides

2- (3-methyl-1-piperidinyl) -6-phenyl-pyrimidine-4-carboxamides

2-azetidin-1-yl-6-phenyl-pyrimidine-4-amides

2- (3, 5-dimethyl-1-piperidinyl) -6-phenyl-pyrimidine-4-carboxamides

2- (butyl- (cyanomethyl) amino) -6-phenyl-pyrimidine-4-carboxamide

6-benzothien-2-yl-2- (1-piperidinyl) pyrimidine-4-amides

2- [ (2-hydroxy-2-phenyl-ethyl) -methyl-amino ] -6-phenyl-pyrimidine-4-carboxamide

6- (2, 4-dichlorophenyl) -2-diethylamino-pyrimidine-4-carboxamide

2- [ [1- (3, 4-dimethoxyphenyl) cyclopropyl ] methylamino ] -6-phenyl-pyrimidine-4-amide

2- (butyl- (2-hydroxyethyl) amino) -6-phenyl-pyrimidine-4-amide

6- (2-methoxyphenyl) -2- (1, 2, 3, 6-tetrahydropyridin-1-yl) pyrimidine-4-amide

6- (3-methoxyphenyl) -2- (1-piperidinyl) pyrimidine-4-amides

N-methyl-6-phenyl-2- (1, 2, 3, 6-tetrahydropyridin-1-yl) pyrimidine-4-amide

2- (cyclopropylmethylamino) -6- (4-methoxyphenyl) -pyrimidine-4-carboxamide

2-diethylamino-6- [4- (trifluoromethyl) phenyl ] -pyrimidine-4-carboxamide

2-diethylamino-6- [4- (trifluoromethoxy) phenyl ] -pyrimidine-4-carboxamide

2- (2-furylmethyl-amino) -N, N-dimethyl-6-phenyl-pyrimidine-4-carboxamide

6-phenyl-2- (1, 2, 3, 6-tetrahydropyridin-1-yl) pyrimidine-4-amide

2- (ethyl- (2-hydroxyethyl) amino) -6-phenyl-pyrimidine-4-amide

2- [2- (1H-indol-3-yl) ethyl-methyl-amino ] -6-phenyl-pyrimidine-4-carboxamide

6- (3-Acetylaminophenyl) -2-diethylamino-pyrimidine-4-carboxamide

2-ethylamino-6-phenyl-pyrimidine-4-amides

2- [ bis (2-hydroxyethyl) amino ] -6- (2-methoxyphenyl) -pyrimidine-4-amide

2-diethylamino-N, N-dimethyl-6-phenyl-pyrimidine-4-carboxamide

2-diethylamino-6- (2, 5-difluorophenyl) -pyrimidine-4-carboxamide

6- (3-methoxyphenyl) -2- (tetrahydrofuran-2-ylmethyl-amino) pyrimidine-4-amide

6- (3, 4-dimethylphenyl) -2- (1-piperidinyl) pyrimidine-4-amides

6-benzo [1, 3] dioxol-5-yl-2- (1-piperidinyl) pyrimidine-4-amides

2-diethylamino-6- (o-tolyl) pyrimidine-4-carboxamide

2- (cyclopropylmethyl-propyl-amino) -6- (4-methoxyphenyl) -pyrimidine-4-amide

2- (isopentyl-methyl-amino) -6-phenyl-pyrimidine-4-amide

2- (isobutyl-methyl-amino) -6-phenyl-pyrimidine-4-carboxamide

2-diethylamino-6- (2, 6-difluorophenyl) -pyrimidine-4-carboxamide

6- (o-tolyl) -2- (1-piperidinyl) pyrimidine-4-carboxamides

44. A method of modulating ABC transporter activity comprising: contacting a cell with a compound of the formula

Wherein

Each Ra is independently H, an optionally substituted aliphatic, an optionally substituted aryl, an optionally substituted aralkyl, an optionally substituted heteroaralkyl, an optionally substituted heteroaryl, an optionally substituted cycloaliphatic, or an optionally substituted cycloheteroaliphatic;

each Rb is independently H, an optionally substituted aliphatic, an optionally substituted aryl, an optionally substituted aralkyl, an optionally substituted heteroaralkyl, an optionally substituted heteroaryl, an optionally substituted cycloaliphatic or an optionally substituted cycloheteroaliphatic orWherein w is 1, 2, 3, 4 or 5, phenyl is optionally substituted with 1-4 Re;

or Ra and Rb, together with the nitrogen atom to which they are bonded, form an optionally substituted heterocycloaliphatic, an optionally substituted heteroaryl, a,

Each Rc is independently H, an optionally substituted aliphatic group, an optionally substituted aryl group, an optionally substituted aralkyl group, an optionally substituted heteroaralkyl group, an optionally substituted heteroaryl group, an optionally substituted cycloaliphatic group, an optionally substituted cycloheteroaliphatic group;

each Rd is independently H, an optionally substituted aliphatic, an optionally substituted aryl, an optionally substituted aralkyl, an optionally substituted heteroaralkyl, an optionally substituted heteroaryl, an optionally substituted cycloaliphatic, an optionally substituted cycloheteroaliphatic, or Rc and Rd taken together with the nitrogen atom to which they are bound form an optionally substituted heterocycloaliphatic;

ring a is aryl or heteroaryl, each optionally substituted with 1-4 Re;

each Re is independently carboxy, amino, nitro, alkylcarbonylamino, cycloalkylcarbonylamino, (cycloalkyl) alkylcarbonylamino, arylcarbonylamino, aralkylcarbonylamino, (heterocycloalkyl) carbonylamino, (heterocycloalkyl) alkylcarbonylamino, heteroarylcarbonylamino, heteroaralkylcarbonylamino, sulfonyl, sulfinyl, sulfanyl, sulfoxy, sulfamoyl, sulfonylamino, ketal, or carbamoyl, cyano, halogen, urea, thiourea, haloalkyl, or-Z-Rf, wherein each alkyl, cycloalkyl, heterocycloalkyl, aryl, and heteroaryl portion of the Re substituent is optionally substituted with 1-3 Rg;

each Z is absent, -O-or-S-;

each Rf is independently hydrogen, alkyl, carboxyalkyl, hydroxyalkyl, alkenyl, alkynyl, cycloalkyl, (cycloalkyl) alkyl, heterocycloalkyl, (heterocycloalkyl) alkyl, aryl, heteroaryl, aralkyl, heteroaralkyl, aroyl, heteroaroyl, alkoxycarbonyl, alkylcarbonyl, aminocarbonyl, or acyl, wherein each alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl, and heteroaryl moiety on the Rf substituent is optionally substituted with 1-3 Rg; while

Each Rg is independently halogen, cyano, alkoxy, hydroxy, nitro, haloalkyl, and alkyl.

45. The method of claim 44, wherein the compound is 2-amino-6- (3-methoxyphenyl) -pyrimidine-4-amide or has the structure of any one of claims 1-43.

46. The method of claim 44, wherein said ABC transporter is CFTR.

47. The method of claim 46, wherein the compound is 2-amino-6- (3-methoxyphenyl) -pyrimidine-4-amide or has the structure of any one of claims 1-43.

48. A pharmaceutical composition comprising 2-amino-6- (3-methoxyphenyl) -pyrimidine-4-carboxamide and a pharmaceutical carrier or a compound according to claims 1 to 43 and a pharmaceutical carrier.

49. A method of treating or lessening the severity of an ABC transporter mediated disease in a mammal comprising the step of administering to said mammal 2-amino-6- (3-methoxyphenyl) -pyrimidine-4-amide or a compound as described in claims 1-43.

50. A method of treating or lessening the severity of a CFTR mediated disease in a mammal comprising the step of administering to said mammal 2-amino-6- (3-methoxyphenyl) -pyrimidine-4-amide or a compound as described in claims 1-43.

51. A method of treating or lessening the severity of cystic fibrosis in a mammal comprising the step of administering to said mammal 2-amino-6- (3-methoxyphenyl) -pyrimidine-4-amide or a compound according to claims 1-43.