Privileged substructures are of potentially great importance in medicinal chemistry. These scaffolds are characterized by their ability to promiscuously bind to a multitude of receptors through a variety of favorable characteristics. This may include presentation of their substituents in a spatially defined manner and perhaps also the ability to directly bind to the receptor itself, as well as exhibiting promising characteristics to aid bioavailability of the overall molecule. It is believed that some privileged substructures achieve this through the mimicry of common protein surface elements that are responsible for binding, such as β- and gamma;-turns. As a result, these structures represent a promising means by which new lead compounds may be identified.

The combinatorial synthesis of bicyclic privileged structures or privileged substructures

Calcium-dependent chloride channels are required for normal electrolyte and fluid secretion, olfactory perception, and neuronal and smooth muscle excitability. The molecular identity of these membrane proteins is still unclear. Treatment of bronchial epithelial cells with interleukin-4 (IL-4) causes increased calcium-dependent chloride channel activity, presumably by regulating expression of the corresponding genes. We performed a global gene expression analysis to identify membrane proteins that are regulated by IL-4. Transfection of epithelial cells with specific small interfering RNA against each of these proteins shows that TMEM16A, a member of a family of putative plasma membrane proteins with unknown function, is associated with calcium-dependent chloride current, as measured with halide-sensitive fluorescent proteins, short-circuit current, and patch-clamp techniques. Our results indicate that TMEM16A is an intrinsic constituent of the calcium-dependent chloride channel. Identification of a previously unknown family of membrane proteins associated with chloride channel function will improve our understanding of chloride transport physiopathology and allow for the development of pharmacological tools useful for basic research and drug development.

http://science.sciencemag.org/content/sci/322/5901/590.full.pdf

TMEM16A, A Membrane Protein Associated with Calcium-Dependent Chloride Channel Activity

Several sporadic and genetic diseases are caused by protein misfolding. These include cystic fibrosis and other devastating diseases of childhood as well as Alzheimer's, Parkinson's and other debilitating maladies of the elderly. A unified view of the molecular and cellular pathogenesis of these conditions has led to the search for chemical chaperones that can slow, arrest or revert disease progression. Molecules are now emerging that link our biophysical insights with our therapeutic aspirations.

Therapeutic approaches to protein-misfolding diseases

Secretory diarrhea is the leading cause of infant death in developing countries and a major cause of morbidity in adults. The cystic fibrosis transmembrane conductance regulator (CFTR) protein is required for fluid secretion in the intestine and airways and, when defective, causes the lethal genetic disease cystic fibrosis. We screened 50,000 chemically diverse compounds for inhibition of cAMP/flavone–stimulated Cl– transport in epithelial cells expressing CFTR. Six CFTR inhibitors of the 2-thioxo-4-thiazolidinone chemical class were identified. The most potent compound discovered by screening of structural analogs, CFTRinh-172, reversibly inhibited CFTR short-circuit current in less than 2 minutes in a voltage-independent manner with KI approximately 300 nM. CFTRinh-172 was nontoxic at high concentrations in cell culture and mouse models. At concentrations fully inhibiting CFTR, CFTRinh-172 did not prevent elevation of cellular cAMP or inhibit non-CFTR Cl– channels, multidrug resistance protein-1 (MDR-1), ATP-sensitive K+ channels, or a series of other transporters. A single intraperitoneal injection of CFTRinh-172 (250 μg/kg) in mice reduced by more than 90% cholera toxin–induced fluid secretion in the small intestine over 6 hours. Thiazolidinone CFTR inhibitors may be useful in developing large-animal models of cystic fibrosis and in reducing intestinal fluid loss in cholera and other secretory diarrheas.

https://dm5migu4zj3pb.cloudfront.net/manuscripts/16000/16112/JCI0216112.pdf

Thiazolidinone CFTR inhibitor identified by high-throughput screening blocks cholera toxin–induced intestinal fluid secretion

The most common cause of cystic fibrosis (CF) is deletion of phenylalanine 508 (ΔF508) in the CF transmembrane conductance regulator (CFTR) chloride channel. The ΔF508 mutation produces defects in folding, stability, and channel gating. To identify small-molecule correctors of defective cellular processing, we assayed iodide flux in ΔF508-CFTR–transfected epithelial cells using a fluorescent halide indicator. Screening of 150,000 chemically diverse compounds and more than 1,500 analogs of active compounds yielded several classes of ΔF508-CFTR correctors (aminoarylthiazoles, quinazolinylaminopyrimidinones, and bisaminomethylbithiazoles) with micromolar potency that produced greater apical membrane chloride current than did low-temperature rescue. Correction was seen within 3–6 hours and persisted for more than 12 hours after washout. Functional correction was correlated with plasma membrane expression of complex-glycosylated ΔF508-CFTR protein. Biochemical studies suggested a mechanism of action involving improved ΔF508-CFTR folding at the ER and stability at the cell surface. The bisaminomethylbithiazoles corrected ΔF508-CFTR in ΔF508/ΔF508 human bronchial epithelia but did not correct a different temperature-sensitive CFTR mutant (P574H-CFTR) or a dopamine receptor mutant. Small-molecule correctors may be useful in the treatment of CF caused by the ΔF508 mutation.

/pdf/small-molecule-correctors-of-defective-df508-cftr-cellular-34ekibgfws.pdf

Small-molecule correctors of defective ΔF508-CFTR cellular processing identified by high-throughput screening

Novel CFTR Chloride Channel Activators Identified by Screening of Combinatorial Libraries Based on Flavone and Benzoquinolizinium Lead Compounds

Benzoflavone activators of the cystic fibrosis transmembrane conductance regulator: Towards a pharmacophore model for the nucleotide-binding domain

Inhibition of the sodium-proton exchanger (NHE) plays an important role in reducing tissue damage during ischemic reperfusion injury; however, pharmacological inhibitors of NHE have restricted access to acutely ischemic tissues because of severely compromised tissue perfusion. We describe the syntheses, characterization, and NHE inhibitory activities of a novel class of amiloride derivatives where peptides are conjugated to the amiloride C(5) amino group. These new peptide-C(5)-amiloride conjugates are inactive; however, peptide residues were chosen such that selective cleavage by neutral endopeptidase 24.11 (enkephalinase) liberates an amino acid-C(5)-amiloride conjugate that inhibits NHE in a glial cell line. These results confirm the feasibility of using peptide-amiloride conjugates as NHE inhibitor prodrugs. We envision the design of analogous peptide-amiloride prodrugs that can be administered prior to ischemic events and subsequently activated by endopeptidases selectively expressed by ischemic tissues.

https://jpet.aspetjournals.org/content/jpet/early/2004/10/27/jpet.104.076984.full.pdf

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Papers

Novel CFTR Chloride Channel Activators Identified by Screening of Combinatorial Libraries Based on Flavone and Benzoquinolizinium Lead Compounds

Benzoflavone activators of the cystic fibrosis transmembrane conductance regulator: Towards a pharmacophore model for the nucleotide-binding domain

Amiloride Peptide Conjugates: Prodrugs for Sodium-Proton Exchange Inhibition

Dioxazocinium Ortho Esters: A Class of Highly pH-Vulnerable Amphiphiles

Synthesis of a C(4)–C(9) eleutheside template from d-glucal