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leky-latky/cftr-protein/zvysuje-cinnost

4,6,4'-trimethylangelicin (TMA) - lumacaftor (VX-809)

  • Psoralen-related compound
  • Potentiates the cAMP/PKA-dependent activation of WT-CFTR
  • Rescues F508del-CFTR-dependent chloride secretion in both primary and secondary airway cells homozygous for the F508del mutation
  • Stabilizes the first membrane-spanning domain (MSD1)
  • Enhances the interface between NBD1 and ICL4 (MSD2)
  • Anti-inflammatory properties
    • Via reduction of IL-8 expression

TMA analogs

  • Generating compounds with no mutagenicity and phototoxicity - TMA analogs
    • Nanomolar concentrations of these analogs significantly rescued F508del CFTR-dependent chloride efflux in FRT-YFP-F508del, HEK-293 and CF bronchial epithelial cells
  • MSD1 was the smallest domain stabilized by TMA analogs, as previously observed for TMA
  • TMA analogs were not effective on F508del-CFTR protein which was already stabilized by a second-site mutation at the NBD1-ICL4 interface.
  • TMA analogs mediate correction by modifying MSD1 and indirectly stabilizing the interface between NBD1 and CL4
  • TMA, like lumacaftor, stabilizes the first membrane-spanning domain (MSD1) of CFTR (Laselva et al., 2016)
  • Enhances the interface between NBD1 and ICL4 (MSD2) (Laselva et al., 2018)
  • Inhibitory effect on Pseudomonas aeruginosa-dependent IL-8 transcription at nanomolar concentrations in CF-derived bronchial epithelial cells (Tamanini et al., 2011)
  • TMA appears to act as a triple-acting compound
    • Anti-inflammatory activity, in addition to corrector and potentiator properties
  • TMA was designated by EMA as an Orphan Drug for the treatment of cystic fibrosis with the code EU/3/13/1137.
  • Has some potential drawbacks
    • Mutagenicity (Bianchi et al., 1990)
    • Photo-reactivity toward DNA pyrimidine bases (Bordin et al., 1991)
      • Not observed when the compound is not directly irradiated with UVA light.
  • www.frontiersin.org/articles/10.3389/fphar.2018.00719/full


5-hydroxytryptamine

AKT activity

  • CFTR activity is largely dependent on AKT activity [19]
  • Dexa is unable to stimulate CFTR activity in the presence of Akt1/2 kinase inhibitor [19]

ATP

  • Opening and closing of CFTR is also dependent on ATP and ADP [17]

Adenosine proteases

Adenoviral infection


Adrenaline (epinephrine)

Alkylxanthine

  • Caffeine, theophylline, theobromine
  • Coffee, tea, cocoa
  • Inhibition of phosphatases
  • Blockade of adenosine receptors [17]
    • Relevant site of action at dietarily achieved alkylxanthine concentrations (50 mcM)
  • Mobilization of intracellular Ca2+
    • 500-1000 mcM concentrations of caffeine or theophylline to increase intracellular Ca2+ or inhibit PDE
  • Inhibition of phophodiesterases
    • To elevate cAMP levels
    • Often intended reason for using alkylxanthines
    • Especially 3-isobutly-1-methyxanthine (IBMX) [17]

IBMX

  • Partially restored amylase and mucin secretion in submandibular acinar cells from CF patients
  • Naturally occurring mutations in the first NBF could be activated by high concentrations of IBMX
  • Mutant CFTR were less sensitive than wild-type CFTR to IBMX
  • Same changes in NBF2 (K1250, Walker A; D1370, Walker B) produced an increase in sensitivity.
  • Missense mutation G480C associated with CFTR protein mislocalization
    • Was equally sensitive to IBMX stimulation when compared with wild-type CFTR
  • IBMX was found to activate del F508 and G551D CFTR-mediated anion efflux



Alkylxanthines

  • Compounds that increase the activity of CFTR [16]

Alpha-glucosidase inhibitors

  • (Norez et al, 2006) miglustat

Antihypertensive 1,4-dihydropyridines (DHPs)

Nifedipine, nicardipine, nimodipine, isradipine, nitrendipine, felodipine, and niguldipine

Antioxidant treatment

Apigenin, kaempferol and genistein

  • Immunofluorescence demonstrated a favorable change in the intracellular distribution of CFTR with 24 h treatments

4-PBA (v.s. možná prozánětlivý)

  • Increase in Cl- conductance as measured by Cl- efflux in cells that were treated for 24 h with 4-PBA
  • Assayed with forskolin and 1 microM or 5 microM genistein
  • Also with cells treated for 24 h with either 4-PBA, 5 microM apigenin, or 1 microM quercetin
    • Combination of chronic treatment with 4-PBA or select flavonoids
      • Followed by acute flavonoid exposure, may be beneficial in cystic fibrosis [7]

Autophagy stimulation could be achieved by:

  • Inhibition of the acetyltransferase EP300
    • Aspirin,
    • Epigallocatechin gallate, EGCG,
    • Spermidine
  • Neutralizing BECN1 inhibitory proteins from the BCL2 family
    • ABT737,
    • Navitoclax,
    • Venetoclax
  • Inhibitors of the mechanistic target of rapamycin complex-1 mTORC1
    • Rapamycin
    • Tacrolimus [25]

Basolateral membrane K+ conductance

  • Utility of augmenting the basolateral membrane K+ conductance
    • In sustaining a Cl- secretory response in the human airway [17]

Benzimidazolone 1-ethyl-2-benzimidazolinone (benzimidazolone 1-EBIO)

  • Stimulated a sustained charybdotoxin-sensitive Cl- secretory response
  • Activates both apical Cl- conductance (CFTR) and basolateral K+ conductance (KCa)
    • Thereby providing the coordinate regulation necessary to stimulate a transepithelial Cl- secretory response
  • Speculate that modulating the driving force on Cl- secretion via the direct activation of a basolateral membrane K+ conductance would be beneficial in modulating Cl- secretion across the human airway [17]

Benzimidazolones - NS004

  • Activated both wild-type and delF508 CFTR
    • Xenopus oocyte expression system
    • Excised outside-out patches from Vero cells transiently expressing the CFTR protein
  • Using I- efflux and whole cell patch-clamp measurements
    • NS004 similarly activated an additional CFTR mutant, P574H
    • Failing to activate the CF-causing R560T or del I507 CFTR mutants
  • NS004 was the first described pharmacological opener of CFTR
  • NS004 activated wild-type CFTR current to 30–60% of that seen with elevated cAMP [17]
  • no effect of NS004 was seen unless CFTR was first activated by cAMP
    • NS004 cannot activate CFTR unless it is in a permissive, phosphorylated state
  • After maximal activation with cAMP, wild-type CFTR cannot be further activated by NS004 in Xenopus oocytes
  • DelF508 CFTR could not be activated in the oocyte expression system unless it was first activated by cAMP [17]
  • After activation of G551D CFTR with a maximal concentration of forskolin (10 µM) plus IBMX (5 mM)
    • Subsequent addition of NS004 (30 µM) induced an additional twofold increase in CFTR-dependent current
  • NS004 and 8-methoxypsoralen
    • Only sporadically activate CFTR in excised inside-out patches
    • Consistently activating the channel in both oocytes and intact epithelial monolayers
    • CFTR must be at least partially phosphorylated before NS004 is capable of activating the channel in Xenopus oocytes
  • NS004 activated an apical membrane Cl- conductance in nystatin-permeabilized T84 monolayers
  • NS004 and NS1619 both further increased Cl- secretion when added subsequent to 1-EBIO and potentiated the response to the Ca2+-mediated agonist carbachol

Benzimidazolones

  • Compounds that increase the activity of CFTR [16]

Benzoxazoles (Chlorzoxazone)

  • Potent openers of basolateral membrane K+ channels
  • Centrally acting muscle relaxants chlorzoxazone and zoxazolamine
    • Activators of KCa in both T84 monolayers as well as primary cultures of HBE
    • Directly stimulated Cl- secretion in these two culture systems
  • 200 µM chlorzoxazone stimulates a sustained Cl- secretory response in our HBE cultures
    • Correlates with the readily obtainable peak plasma concentrations found in vivo
  • Both chlorzoxazone and zoxazolamine
    • Activated KCa in excised inside-out patches
  • Chlorzoxazone induces a hyperpolarization of the transepithelial nasal potential difference in normal volunteers
    • After inhibition of the basal Na+ absorption with amiloride
    • Hyperpolarization is indicative of an induced Cl- secretory response
  • Chlorzoxazone is approved by the Food and Drug Administration
    • Therapy for the relief of acute musculoskeletal pain
      • Benzodiazepines have largely displaced the use of these muscle relaxants
    • High doses (600–750 mg)
      • Peak plasma concentration of 100–200 µM is obtained
      • Half-life of 1 h
    • Concentration in kidney and muscle is approximately one-half that found in plasma
  • Preferentially suppress polysynaptic reflexes
  • Decrease striatal dopamine turnover
  • Decrease neuronal firing rate
  • Does not appear to involve strychnine-insensitive glycine or GABAB receptors
  • Direct effect on KCa
    • Direct effect on an ion-conductive pathway may be important in determining their clinical efficacy.
  • SK channels have a principally neuronal expression pattern
  • IK is expressed peripherally in epithelial tissues including lung and colon
    • RSK2 and hIK1
  • Chlorzoxazone, 1-EBIO, and zoxazolamine
    • All activate both hIK1 and rSK2
      • Dependent on the presence of physiological levels of cytoplasmic Ca2+
      • Predicted to hyperpolarize the neuron thus decreasing firing rate as observed
  • Metabolic product of chlorzoxazone is 6-hydroxychlorzoxazone [17]

Beta2 mimetika

  • Cystic fibrosis (CF) s CFTR gene mutations associated with residual function
    • May benefit from G-protein coupled receptor (GPCR)-targeting drugs that can activate and enhance CFTR function
  • Beta 2-adrenergic receptor agonists as the most potent inducers of CFTR function
  • Beta 2-Agonist-induced could induce in homozygous CFTR-F508del after rescue of CFTR trafficking by small molecules
  • In vivo response to oral or inhaled ß2-agonist (salbutamol) in CF patients with residual CFTR function was evaluated in a pilot study
    • 10 subjects with a R117H or A455E mutation
    • Showed changes in the nasal potential difference measurement after treatment with oral salbutamol
      • Significant improvement of the baseline potential difference of the nasal mucosa (+6.35 mV, p<0.05)
      • Plasma that was collected after oral salbutamol treatment induced CFTR activation when administered ex vivo to organoids
  • pubmed.ncbi.nlm.nih.gov/27471203/
  • Transforming growth factor-beta 1 and cigarette smoke inhibit the ability of ß2-agonists to enhance epithelial permeability.
  • Beta-adrenergic bronchodilators (such as albuterol), is nearly universal for airway clearance
  • Greater than 60% reduction in both wild-type and modulator-corrected F508del-CFTR activation following chronic exposure to short- and long-acting ß-agonists !!!
  • insight.jci.org/articles/view/93029
  • CFTR activation via the beta-2 adrenergic receptor (beta2AR)
    • C-terminal PDZ binding domains (present in both proteins)
    • Interact with G proteins (GPs)
      • Dissociation of GSalpha
        • Which activates adenylyl cyclase (AC)
          • Catalyzes the conversion of ATP to cAMP
    • A-kinase anchoring proteins help localize other critical proteins
        • AC,
        • Phosphodiesterases [PDEs]
        • Multidrug resistance protein 4 [MRP4]
      • That titrate and compartmentalize cAMP levels and subsequent CFTR activity
  • insight.jci.org/articles/view/93029
  • Often the short acting salbutamol or the long acting salmeterol is used by inhalation.
  • Two month double-blind crossover trial of 90 µg salbutamol four times daily
    • Significantly improved peak expiratory flow rate (PEFR) in patients with bronchial hyperresponsiveness
    • Lung function was not changed in this study
  • Salbutamol (pMDI, 180 µg b.i.d.) (Ventolin)
    • Significantly improved respiratory function in a 12 month observational study
  • Placebo-controlled double-blinded trial, CF patients
    • Six months of 180 µg inhaled salbutamol twice daily
      • Did not differ significantly compared to placebo in lung function tests
  • Study 18% of CF patients, who had salbutamol
    • Significant increase in FEV1
  • Inhaled short-acting beta-agonists
    • Did not improve exercise performance or post exercise dyspnoea in CF patients despite significantly improving FEV1

  • Stable CF patients who responded to day time salbutamol
    • Showed significant increases in nocturnal oxyhaemoglobin saturation
      • Following salmeterol administration before sleep
  • A Cochrane review concluded that both short and long acting ß-sympathomimetics
    • Can be beneficial in CF patients with bronchodilator responsiveness or bronchial hyperresponsiveness
  • Bronchial smooth muscle relaxation may increase airway compression and reduce cough efficiency by inducing large airway collapse
    • Negative responses are unusual and collapse is unlikely during normal breathing
    • During exacerbations, the efficacy of inhaled bronchodilator therapy may be reduced
      • Has not been confirmed in later studies with inhaled salbutamol and high dose salmeterol
  • Combinations of beta-sympathomimetic and anticholinergic drugs
    • Did not result in synergistic or additive effects in CF patients

Long acting bronchodilator salmeterol (AIRFLUSAN, ASTHMEX, AURASTH, BRECUR AIRMASTER )

  • Unblinded study dyspnoea improved even in patients not showing a positive FEV1 response
  • Treated with 50 µg salmeterol twice daily for two weeks
  • 24 week treatment period, 100 µg salmeterol given twice daily
    • Was well tolerated
    • Associated with better pulmonary function
    • Fewer interventions
    • Fewer respiratory symptoms compared to treatment with salbutamol in CF patients with mild to moderate disease
  • www.sciencedirect.com/science/article/pii/S1569199309000356

Beta-2mimetika s krátkým účinkem (SABA)

  • Salbutamol,
  • Terbutalin
  • Fenoterol

Beta2mimetika s dlouhým až 12-hodinovým trváním účinku (LABA)

  • Formoterol
  • Klenbuterol p.o.

Pomalý nástup účinku a krátké trvání

  • P.o. salbutamol

Pomalý nástup účinku a dlouhé trvání

  • Inhalační salmeterol

NÚ beta2mimetik

  • Tachykardie,
  • Zvýšení krevního tlaku
  • Třes
  • Neměla by být proto podávány u pacientů s kardiovaskulárním rizikem

Ca2+-mobilizing secretagogues


Ca2+

  • CFTR is notstimulated directly by Ca2+
  • Several adenylyl cyclase iso-forms are Ca2+ activated
    • Types 1, 8 and perhaps 3 (Halls&Cooper, 2011) [20]
  • Thus could couple the phosphorylationof CFTR by PKA to muscarinic Ca2+ responses [20]
  • Regulation may involve a localized pool of cAMP near the plasma membrane (Monterisiet al.2012)
    • Isolated from the bulk cytoplasm by phosphodiesterases 4D (Barneset al.2005) and/or 3A (Penmatsuet al.2010) [20]
  • Ca2+ activates CFTR, at least in part
    • Through activationof the Pyk2/Src complex and tyrosine phosphorylation [20]
  • Ca2+ - dependent activation of Src (presumably through its association with Pyk2)
    • Can enhance channel activity by inhibiting the dephosphorylation of PKA sites on CFTR [20]
  • Calcium mobilization is necessary for CFTR activation during muscarinic stimulation [20]

Caffeine (1, 3,7-trimethylxanthine)

  • Promoted Cl- secretion through enterocyte apical CFTR
    • Thus inhibited sodium absorption
  • Depletion of cAMP or inhibition of CFTR
    • Completely abolished the effect of caffeine on Cl- secretion
  • Chronic caffeine consumption reduces sodium absorption by promoting CFTR-mediated Cl- secretion in the intestine
    • Which contributes to the anti-hypertensive effect of caffeine in salt-sensitive rats. [18]
  • Rapidly absorbed from the gastrointestinal tract and eliminated in 4-6 h
  • Acute caffeine intake can raise blood pressure by increasing the activity of the sympathetic nervous system
  • Chronic use lowers blood pressure by promoting renal sodium excretion
  • Associated with increase in intracellular cAMP levels
    • By caffeine-induced activation of ryanodine receptors
    • Leads to Ca2+ release from the Ca2+ store
      • Inhibition of phosphodiesterase
        • Increase in the level of intracellular cAMP
          • Could also regulate the activity of NHE3 or CFTR [18]

Carbachol (Miostat)

  • choline carbamate
  • Positively charged quaternary ammonium compound
  • Parasympathomimetic cholinergic agonist
  • Mimics the effect of acetylcholine on both the muscarinic and nicotinic receptors [2]
  • Carbachol canamplify the response to VIP
    • By stimulating
      • Ca2+- activated high conductance
      • Intermediate conductance K+channels
    • Thereby hyperpolarizing the cell membrane and increasing the driving force for apical anion efflux [20]
  • CFTR can also be activated by muscarinic agonist carbachol (CCh) - stimulates:
    • Phospholipase C,
    • Ca2+ mobilization
    • Src tyrosine kinase regulation (Billet and Hanrahan, 2013; Billet et al., 2013) [22]

VIP and muscarinic agonist carbachol

  • Increase CFTR aggregation into cholesterol-dependent clusters
  • Reduce CFTR lateral mobility within and between membrane microdomains
  • Trigger the fusion of clusters into large (3.0 µm2) ceramide-rich platforms
  • CFTR clusters are closely associated with
    • Motile cilia
    • Enzyme acid sphingomyelinase (ASMase)
      • Constitutively bound on the cell surface [22]
  • Formation of ceramide-rich platforms containing CFTR
    • Enhances transepithelial secretion
    • Likely has other functions related to inflammation and mucosal immunity.
  • Platforms are reversible
  • Induction does not lead to an increase in apoptosis
  • Platform induction is prevented by pretreating cells with
    • cholesterol oxidase to disrupt lipid rafts
    • Exposure to the ASMase functional inhibitor amitriptyline
    • Membrane-impermeant reducing agent 2-mercaptoethanesulfonate [22]
  • Blocking platform formation
    • Does prevent the increase in CFTR surface expression that normally occurs during VIP stimulation [22]

Oční aplikace

  • Administered ocularly to induce miosis
    • To reduce intraocular pressure in the treatment of glaucoma [2]
    • Decrease intraocular pressure in people with primary open-angle glaucoma [2]
    • Long-term therapy of non-congestive wide-angle glaucoma
  • Principal effects are miosis and increased aqueous humour outflow
  • Use during ophthalmic surgery
    • Sometimes used to constrict the pupils during cataract surgery [2]
  • Carbachol is also used to stimulate micturition by contraction of detrusor muscle
  • Not well absorbed in the gastro-intestinal tract
  • Does not cross the blood–brain barrier
  • Not easily metabolized by cholinesterase
  • 2 to 5 minute onset of action
  • Duration of action is 4 to 8 hours with topical administration
  • 24 hours of action for intraocular administration
  • Benzalkonium chloride is mixed in to promote absorption [2]
  • Carbachol eyedrops are used to decrease the pressure in the eye for people with glaucoma

Nosní sliznice

Co by se stalo, kdyby si je kapali do oteklého nosu lidé u kterých nefunguje Nasivin (třeba při COVID-19?)

GIT

  • Strong promoter of ICC activity, which is mediated through the calcium-activated chloride channel, anoctamin
  • Carbachol-induced increase in membrane current was dependent on NKCC1 co-transport, basolateral K(+) channels and Cftr activity
  • Increase in capacitance was partially dependent on NKCC1 and K(+) channel activity, but did not require Cftr activity
  • Carbachol also induced an increase in mucus thickness that was inhibited by the NKCC1 blocker bumetanide
  • Mice that lacked a functional Cftr channel did not respond to carbachol with an increase in mucus thickness
    • Carbachol-induced mucin expansion requires Cftr channel activity [4]
  • Carbachol-stimulated HCO3-secretion pancreatic
    • Reduced by75–100% in the intestine ofcftrnull mice (Hoganet al.1997; Seidleret al.1997) [20]

CNS zvířat

  • Cat and rat
    • Induce rapid eye movement (REM) sleep when microinjected into the pontine reticular formation [2]
    • Via activation of postsynaptic muscarinic cholinergic receptors (mAChRs) [2]

Močová retence

  • Stimulate bladder emptying if the normal emptying mechanism is not working properly.
  • Oral (2 mg) tablet [2]

Srdce

  • Chronic Treatment With Carbachol (4 day infusions) Sensitizes the Myocardium to cAMP-Induced Arrhythmia
  • Changes were accompanied by a
    • Decrease in left ventricular M-cholinoceptor density by 15% (P<.05)
    • Decrease in pertussis toxin–sensitive G proteins (Gi?) by 26% (P<.05) without a decrease in the corresponding mRNAs.
    • Beta-Adrenoceptor density and basal and stimulated adenylyl cyclase activity remained unchanged [3]

Průmysl

  • Classified as an extremely hazardous substance in the United States as defined in Section 302 of the U.S. Emergency Planning and Community Right-to-Know Act (42 U.S.C. 11002) [2]

Kontraidikace

  • Asthma
    • Nastane bronchokonstrikce nebo zhoustne / zmnoží se / nabobtná do té doby hustý hlen ?
  • Coronary insufficiency,
  • Gastroduodenal ulcers,
  • Incontinence
  • Drug will exacerbate the symptoms of these disorders [2]


Chenodeoxycholic acid (CDCA) and deoxycholate

  • Activate rabbit colonic mucosal adenylate cyclase activity
  • Decrease phosphodiesterase activity
  • Hence raise intracellular cAMP
    • Thereby stimulating colonic fluid secretion
  • When CFTR is activated, it raises Cl- conductance and together with activated K+ conductances permits net NaCl and water secretion into the luminal solution, since intracellular Cl- activity is raised by basolateral Na+K+Cl-2 cotransporter activity.
  • facultyopinions.com/prime/736126318

Cholera

Cholinergic stimuli

  • CFTR also responds to cholinergic and purinergic responses in native tissues
  • CFTR itself mediates a substantial fraction ofthe ‘CaCC conductance’ during muscarinic stimulation of airway submucosal glands
    • CFTR activation results from Ca2+- dependent phosphorylation by both PKA and tyrosine kinases. [20]
  • Co-expressingCFTR together with the type 3 muscarinic receptor (M3R)in baby hamster kidney (BHK) cells (Billetet al.2013) [20]
  • CFTR currents stimulated by carbachol in heterologous expression system were about 40% largerthan those induced by forskolin [20]
  • Cholinergic stimulation of airway epithelial cells
    • By the M3R and, to a lesser extent, the M1 receptor
  • M3R signals - via G alpha q/11 to phospholipase C
    • Generates IP3
    • Mobilizes Ca2+ from intracellular stores (Berridge, 2012) [20]
  • physoc.onlinelibrary.wiley.com/doi/pdf/10.1113/jphysiol.2013.261909


Correct the trafficking and functional defects of the del.F508 mutant

  • Correct the DF508-CFTR trafficking defect:
    • Aminoarylthiazoles,
    • Quinazolinylaminopyrimidinones,
    • Bisaminomethylbithiazoles
      • Corr-4a (Pedemonte et al, 2005b)
    • 1,4-dihydropiridines (Pedemonte et al, 2005a)
    • Quinazolines
      • VRT-325 (Loo et al, 2005)
    • Sildenafil analogues
      • KM11060 (Robert et al, 2008)
      • Galfenine (Robert et al, 2010)
    • VX-809 (Van Goor et al, 2011) - most potent
  • VRT-325 and VRT-532
    • May directly interact with DF508- NBD1 and thermally stabilize the protein.
    • VRT-325 exhibits undesirable effect
      • Inhibits the ATPase activity of DF508-CFTR by decreasing its affinity for ATP (Kim et al, 2010).
    • [EMBO Mol Med (2013) 5, 1484–1501, 2013 The Authors. Published by John Wiley and Sons, Ltd on behalf of EMBO, Research Article Norbert Odolczyk et al.]

Correctors

Class 1

N-terminal domain of CFTR (i.e., MSD1) was important for the stabilizing effect

  • Of lumacaftor (Loo et al., 2013; Ren et al., 2013; Laselva et al., 2016).
  • Full-length mutant protein F508del-CFTR
    • A secondary “rescue” mutation that partially compensates for defective interaction of NBD1 and ICL4, namely R1070W
    • Partially abrogated the correction mediated by lumacaftor
      • Lumacaftor mediates its correction via allosteric effects on this interface (He et al., 2013; Okiyoneda et al., 2013; Laselva et al., 2018)
  • www.frontiersin.org/articles/10.3389/fphar.2018.00719/full

Curcumin

  • These compounds appear to function as allosteric modulators that circumvent ATP binding defects by promoting ATP-free channel activity
  • W1282X-CFTR-mediated currents across FRT monolayers that were stimulated by these compounds were small relative to wild type CFTR (ca 5%)
  • curcumin exerted additive effects on W1282X-CFTR channel gating (opening/closing)
    • Combinations of CFTR potentiators of different mechanistic classes to activate the W1282X-CFTR channel might be therapeutically beneficial
  • Robustly stimulate del 1198-CFTR channel activity
  • 30 microM curcumin and 300 nM VX-770
    • Had additive effects on the macroscopic currents mediated W1282X-CFTR
  • Curcumin strongly increased the current mediated by this construct well above the level stimulated by a maximally effective dose of VX-770
  • journals.plos.org/plosone/article?id=10.1371/journal.pone.0152232
  • curcumin may act by modifying keratin 18 (K18) network (Hamdaoui et al, 2011; Lipecka et al, 2006)
  • K18 heterodimerizes with keratin 8 (K8)
  • Role in the trafficking of CFTR/ DF508-CFTR (Colas et al, 2012; Duan et al, 2012).

Cysteamine

  • A TGM2 inhibitor that prevents TGM2-mediated BECN1 sequestration
  • Can reestablish autophagic flux and restore the function of the F508del-CFTR mutant at the epithelial surface
  • Both in patients and in mice models bearing a similar CFTR mutation
  • Beneficial effects of cysteamine on both CFTR function and autophagy persist for several weeks after cysteamine withdrawal
  • Restoration of proteostasis results in transient homeostasis before the system again loses its balance [25]
  • By the degradation of coenzyme A
  • Biosynthetic precursor to the neurotransmitter hypotaurine
  • Stable aminothiol
  • Organic compound containing both an amine and a thiol functional groups
  • Often used as salts of the ammonium derivative [HSCH2CH2NH3]+[12] including the hydrochloride, phosphocysteamine, and bitartrate
  • As a medication, cysteamine - Cystagon among others
    • Indicated to treat cystinosis by mouth (capsule and extended release capsule) and in eye drops
  • Warnings about symptoms similar to
    • Ehlers-Danlos syndrome,
    • Severe skin rashes,
    • Ulcers or bleeding in the stomach and intestines
    • Central nervous symptoms including seizures, lethargy, somnolence, depression, and encephalopathy
    • Low white blood cell levels,
    • Elevated alkaline phosphatase,
    • Idiopathic intracranial hypertension
      • Can cause headache, tinnitus, dizziness, nausea, double or blurry vision, loss of vision,
      • Pain behind the eye or pain with eye movement
  • Additional adverse effects of oral cysteamine include
    • Bad breath,
    • Skin odor,
    • Vomiting, nausea, stomach pain, diarrhea, and loss of appetite
  • For eye drops, the most common adverse effects are
    • Sensitivity to light, redness, and eye pain, headache, and visual field defects
  • People with cystinosis lack a functioning transporter (cystinosin) which transports cystine from the lysosome to the cytosol
    • Leads to buildup of cystine in lysosomes, where it crystallizes and damages cells
  • Cysteamine enters lysosomes and converts cystine into
    • Cysteine
    • Cysteine-cysteamine mixed disulfide
      • Both of which can exit the lysosome
  • Cysteamine promotes the transport of L-cysteine into cells
    • Can be further used to synthesize glutathione
  • In in vitro and animal models for radiation protection in the 1950s
  • For sickle cell anemia
  • Effects on growth
  • Ability to modulate the immune system
  • As a possible inhibitor of HIV
  • An antioxidant with chemo-sensitizing and radioprotective properties

Decreased NEDD4L protein function

  • Vlivem glukokortikoidů [18]
  • Na+ transport is strongly affected by NEDD4L and phosphorylation of NEDD4L
    • Thus increases epithelial Na+ channel (ENaC) membrane abundance by preventing its endocytic retrieval [18]


Dexamethason

  • Elevate CFTR activity in airway epithelial cells
  • Phosphoinositide 3-kinase (PI3K) pathway is involved in CFTR stimulation
  • Critical involvement of PI3K and protein kinase B (AKT) signaling in the dexa-induced increase of CFTR activity
  • Serum and glucocorticoid dependent kinase 1 (SGK1) activity was not essential
  • Increases of CFTR activity by dexa were demonstrated within 30 min accompanied by rapid activation of AKT
  • Dexa induces a rapid stimulation of CFTR activity
    • Depends on PI3K/AKT signaling in airway epithelial cells
  • Glucocorticoids might thus represent, in addition to their immunomodulatory actions, a therapeutic strategy to rapidly increase airway fluid secretion. [19]
  • Effect of hormones on CFTR function, which appears to differ between the distal and the proximal airways
    • GCs reduced CFTR expression and activity in distal lung epithelial cells
      • Responsible for gas exchange and fluid absorption
    • GCs stimulated CFTR activity in the air conducting proximal airways
      • Which are responsible for MCC
      • Its gene expression was also reduced, yet not as pronounced as in the distal epithelia [19]
  • GCs increased CFTR protein expression two-fold
    • Attributed to an altered chaperone interaction resulting in increased CFTR protein trafficking [19]
  • In the transformed cystic fibrosis (CF)
    • Bronchial epithelial cell line CFBE41o-
      • GC-induced increase of serum and GC dependent kinase 1 (SGK1) protein
      • SGK1 phosphorylates and thereby inhibits the ubiquitin ligase neural precursor cell expressed,
        • Developmentally down-regulated protein 4-like (NEDD4L),
        • A mechanism possibly responsible for the elevated CFTR protein abundance [19]
  • Kinase signaling is majorly involved in stimulated CFTR activity induced by GCs in airway epithelial cells [19]
  • PI3K pathway is involved in CFTR stimulation by glukokort. [19]

Within the proximal air-conducting compartment

  • GCs promote fluid secretion
    • By enhancing CFTR activity as shown in Calu-3 and primary airway epithelial cells
  • Airway fluid secretion enhances MCC by maintaining the airway surface liquid layer
  • Dexa increased CFTR protein expression two-fold
    • Attributed to an altered chaperone interaction resulting in increased CFTR protein trafficking, while CFTR gene expression was also reduced

In distal airways

  • The absorptive phenotype predominates
    • Enabling gas exchange in the alveoli
  • GCs
    • Reduced alveolar CFTR activity in distal lung cells
    • Whereas alveolar ENaC activity was increased [19::
    • Promoting fluid absorption to ensure efficient gas exchange
  • GCs reduced CFTR gene expression in distal, but also in proximal lung compartments
  • Stimulation of CFTR activity in airway epithelial cells is most likely mediated by non-genomic mechanisms [19]

Activation of receptor tyrosine kinases results in activation of PI3K

  • Catalytic domain of PI3K subsequently converts phosphatidylinositol (3,4)-biphosphate (PIP2) to phosphatidylinositol (3,4,5)-triphosphate (PIP3),
    • Leading to activation of
      • Phosphoinositide-dependent protein kinase (PDK1)
      • Mammalian target of rapamycin complex 2 (mTORC2)
  • Followed by phosphorylation and activation of AKT
  • Inhibition of either the GR or the PI3K and knock-down of SGK1 blocks the effect of dexa on CFTR trafficking [18]

Diabetics with Sympathetic Nervous Dysfunction

  • Increased Lung Uptake of Iodine-123-MIBG
  • Lung uptake of 123I-MIBG at 4 hr was significantly increased in the diabetic group as compared with others
  • In diabetic patients with sympathetic nervous dysfunction
    • Lung uptake ratio of 123I-MIBG at 4 hr was significantly higher than that in the diabetic patients without sympathetic nervous dysfunction
  • Lung uptake of 123I-MIBG was increased and lung washout of 123I-MIBGwas decreased in diabetic patients with sympathetic nervous dysfunction
  • Lung uptake of 201TIwas not altered.
  • Metaiodobenzylguanidine (MIBG) is an analog of the adrenergic neuron-blocking agent guanethidine
    • Shares the same uptake, storage and release mechanisms as norepinephrine in sympathetic nerve endings
    • Myocardium is richly supplied with sympathetic nerves
      • Myocardial imaging using I23I-MIBG to assess cardiac sympathetic nervous activity
  • MIBG is taken up by the same sodium-dependent, energy-requiring transport system used for the extraction of norepinephrine in pulmonary capillary endothelial cells.
    • [Shuichi Murashima, Kan Takeda, Kaname Matsumura, Koichiro Yamakado, Hajime Sakuma, Tokio Kitano, Tsuyoshi, Nakagawa, Takashi Ichihara, Tetsu Yamakado and Kazuya Murata; Department of Radiology, the First Department of Internal Medicine and the Third Department of Internal Medicine, Mie; University School of Medicine, Mie; Toshiba, Tochigi, Japan ]

Epigallocatechin-gallate (EGCG)

  • CF patients bearing class II CFTR mutations
  • Inhibitor of the autophagy-inhibitory acetyl transferase EP300
  • Can prolong the beneficial effects of cysteamine
    • With respect to autophagy induction
    • Restore CFTR function in nasal respiratory epithelial cells
  • Preclinical studies involving F508del-CFTR mice
    • Cysteamine plus EGCG
      • Loses its capacity to restore CFTR function in a Becn1 haploinsufficient (Becn1+/-) background
        • Autophagy is required for sustaining a functional CFTR at the cell surface [25]

Estrogen

  • Up-regulace exprese genu CFTR

Flavones

  • Compounds that increase the activity of CFTR [16]

Fluoride

  • F- activates CFTR activity in the cell-attached configuration
    • Stably active CFTR channels in excised membrane patches are unaffected by the presence of 10 meq F- [17]
  • Affects a variety of hydrolytic enzymes including
    • G proteins,
    • Kinases,
    • Phosphatases [17]
  • Concentration dependence for specific enzymes ranges from low microequivalent to the milliequivalent range
  • 10 meq F- blocked the effect of exogenously added alkaline phosphatase
    • Channel rundown was inhibited by this treatment [17]
  • 20 meq F-
    • Increased CFTR Po 2x [17]
  • 5 milliequivalents F-
    • Slightly reduced, but did not inhibit, the reduction in Cl- conductance that accompanies the washout of cAMP from a permeabilized human sweat duct

Forskolin

  • Product of Coleus forskohlii
  • Binds directly to the catalytic subunit of ACs
    • Dimerizing the C1 and C2 domains
      • Activating the enzyme independently of GP activity
  • Regulation of AC function is unique to each isoform
  • insight.jci.org/articles/view/93029
  • Forskolin (an adenylate cyclase stimulator) to maximally activate the CFTR channel

CFTR Phosphorylation is Needed for its Channel Activity

  • CFTR channel function depends on the conformational changes driven by ATP binding and hydrolysis at the heterodimer NBD1:NBD2
    • Shifting the molecule between the open and closed states (Anderson et al., 1991)
  • Transition in conformation is initiated by protein kinase A (PKA)-dependent phosphorylation of the R domain
  • The open probability of the channel is dependent on R domain phosphorylation at multiple sites
    • Reflecting a balance between protein kinase and phosphatase activity (Cheng et al., 1991)
  • CFTR is primarily phosphorylated by PKA
  • www.frontiersin.org/articles/10.3389/fchem.2016.00001/full
  • In phosphorylated CFTR channels opening and closing (gating) of the anion pore is coupled to
    • Conformational changes induced by ATP binding and hydrolysis at two cytosolic nucleotide binding domains (NBDs)
      • (Anderson et al., 1991; Li et al., 1996).
  • elifesciences.org/articles/46450

G protein-mediated activation of adenylyl cyclase


Genistein

  • Reported to stimulate CFTR by modulating phosphatase activity [17]
  • Plant-derived isoflavone
  • Thought to be the estrogen receptor
  • Genistein stimulates Na+ absorption in human CF airway [17]
  • Genistein inhibited tyrosine kinase activity of the
    • Epidermal growth factor (EGF) receptor
    • Src kinase with IC50 values of 2.6 and 30 mcM
    • Having no effect on PKA at 375 µM
  • Genistein was the most cytotoxic to Rous sarcoma virus-transformed cells (26 µM)
    • Genistein (i) nad kaempferol (f) = prunetin (i) = quercetin (f) nad apigenin (f) = biochanin A (i) = acacetin (f) = flavone (f) nad genistin (i) = daidzein (i) [17]
    • Inhibition of EGF receptor tyrosine kinase activity by genistein is
      • Competitive with ATP
      • Noncompetitive or mixed inhibition with the phosphate acceptor
    • Inducer of DNA strand breakage due to its interaction with topoisomerase II (effective concentration 5 mcM)
    • Inhibit PKC and PKA
    • Inhibiting beta-galactosidase [17]
  • Genistein is the most potent of many flavones and isoflavones that have been reported to inhibit
    • protein tyrosine kinases
    • Topoisomerase
    • Certain other ATP-requiring enzymes [17]
  • Genistein and some related flavones and isoflavones affect epithelial ion transport
  • Genistein inhibits dephosphorylation of CFTR resulting in increased and prolonged channel activity [17]
  • Genistein affects CFTR-mediated ion transport by
    • Direct interaction with the channel
    • By inhibition of protein phosphatases
    • By inhibition of tyrosine kinases [17]
  • 125I efflux and cell-attached membrane patches, the concentration of genistein employed (40–50 mcM) [17]
  • Daidzein
    • Analog known not to affect EGF receptor tyrosine kinase
    • Was without effect [17]
  • Vanadate
    • Known tyrosine phosphatase inhibitor inhibited the response [17]
  • CAMP, Ca2+ were not affected by genistein
  • Largely supported by Sears et al.
    • Genistein did not affect intracellular cAMP or Ca2+ concentrations [17]
  • Genistin
    • Analog of genistein that does not affect EGF tyrosine kinase activity was without effect [17]
  • Genistein potentiated, by fourfold, the effect of forskolin (10 mcM) on intracellular cAMP production in T84 cell monolayers
  • Genistein, but not genistin, stimulated bumetanide-inhibitable Cl- secretion in shark rectal glands
    • Tissue was less sensitive to basolateral genistein application
  • 50 mcM genistein prevented carbachol-induced phosphorylation of three proteins
    • Inhibited carbachol-induced Ca2+ entry [17]
  • Other known tyrosine kinase inhibitors
    • Methyl-2,5-dihydroxycinnamate
    • Lavendustin A
    • But not daidzein [17]
  • Genistein-stimulated Cl- currents in whole cell records of guinea pig cardiac myocytes
    • Stimulation was vanadate sensitive
    • Daidzein was virtually without effect [17]
  • Genistein
    • Could, by inhibiting a phosphatase, increase the functional activity of CFTR [17]
  • CFTR is similarly phosphorylated in the presence of either forskolin or genistein
  • Genistein potentiates the effect of maximally effective concentrations of IBMX (5 mM) and forskolin (10 mcM)
    • On G551D CFTR-expressing oocytes
    • no effect on unstimulated or maximally stimulated oocytes expressing wild-type CFTR [17]

Histone deacetylase (HDAC) inhibitors (HDACi)

  • Can have significant benefit in correcting protein-misfolding diseases
  • Cystic fibrosis (CF) is a familial autosomal recessive disease
  • Potential utility of HDACi in correcting the phenylalanine 508 deletion (F508del) CFTR variant remains controversial
  • Provide functional correction of Class II and III CFTR variants
  • Restoring cell surface chloride channel activity in primary human bronchial epithelial cells. We further demonstrate a s
  • Synergistic effect of these HDACi with Vx809
    • Can significantly restore channel activity for multiple CFTR variants
  • HDACi can serve for correcting CF-causing mutations

Histone deacetylase-7 inhibitors

  • SAHA -(Hutt et al, 2010)
  • Interact with proteins responsible for DF508-CFTR processing
    • Increase the amount of DF508-CFTR at the plasma membrane

Panobinostat (LBH-589)

Romidepsin (FK-228)


Imperatorin

  • Del.F508-CFTR including [19]

Inhibition of MAPK

Inhibition of TGM2 and the restoration of autophagy

  • Re-establishes CFTR function at the cell surface [25]

Inhibitors of GSK-3beta

Inhibitors of the Ras/Raf/MEK/ERK or p38 pathways


Inhibitors of receptor Tyr kinases (RTKs)

Isobutylmethylxanthine (Fsk/IBMX)


Isoflavones

  • Compounds that increase the activity of CFTR [16]

Isoprenaline

  • Stimulation by isoprenaline (CFTR-dependent component)
  • In wild-type mice, isoprenaline increased secretion by more than half proti CF myši [14]

Isoproterenol

  • Beta adrenergic agonist
  • Stimulates amiloride-insensitive short-circuit current (Isc)
    • Across non-CF but not CF cell monolayers lacking functional CFTR (Widdicombe et al., 1985)

Isopsoralen

  • Del.F508-CFTR including [19]

Ivacaftor ( Kalydeco, VX-770)

  • Approved by the FDA in 2012 for people with cystic fibrosis
  • Developed by Vertex Pharmaceuticals in conjunction with the Cystic Fibrosis Foundation
  • First drug that treats the underlying cause rather than the symptoms of the disease
  • One of the most expensive drugs, costing over US$300,000 per year, which has led to criticism of Vertex for the high cost.
  • Small-size potentiator approved for treatment of patients carrying the CFTR class III mutation G551D
    • Along with nine other rare ‘gating’ defect mutations (Van Goor et al., 2009, 2014; Yu et al., 2012).
  • Thousands of patients are being treated with the drug
  • Strong, but greatly delayed, channel activation by picomolar Vx-770 identifies multiple sequential slow steps in the activation pathway
  • elifesciences.org/articles/46450
  • By Vertex Pharmaceuticals
    • Using high-throughput screening
  • Patients carrying G551D and other gating mutations (Ramsey et al., 2011)
    • Pod 5% of all CF cases
  • elifesciences.org/articles/46450

  • Del F508 also experienced significant clinical improvement from co-administration of Vx-770 with a combination of corrector drugs (Boyle et al., 2014; Wainwright et al., 2015; Davies et al., 2018).
  • elifesciences.org/articles/46450
  • CFTR channel stimulation by Vx-770 in cell-free patches is reportedly irreversible
    • (Jih and Hwang, 2013; Lin et al., 2016; Yeh et al., 2015; Yeh et al., 2017; Wang et al., 2014; Wang et al., 2018)
  • CFTR Channel Stimulation by Vx-770 is fully reversible
  • Vx-770 stimulates WT and mutant CFTR channels already at subnanomolar concentrations
  • elifesciences.org/articles/46450
  • Prolonged treatment with the potentiator ivacaftor
    • Induces a dose-dependent reversal of the lumacaftor-mediated F508del-CFTR rescue in human bronchial epithelial cultures
    • An increased turnover rate at the cell surface of rescued F508del-CFTR resulting decreased stability
      • Proposed as the underlying mechanisms (Cholon et al., 2014; Veit et al., 2014)
  • www.frontiersin.org/articles/10.3389/fphar.2018.00719/full

LMTK2 knockdown

  • Increased CFTR mediated Cl- secretion in cells expressing wt-CFTR (Luz et al., 2014)
  • Cells expressing F508del-CFTR too
    • Increase in the amount of corrector VX-809-rescued F508del-CFTR (mature band C)
  • Differentially affected the biochemical and functional rescue of F508del-CFTR
    • 10% increase in CFTR band C abundance translated into a 50% rise of the forskolin/IBMX-stimulated short-circuit current.
  • Clinical studies have already shown that targeted monotherapy with the CFTR correctors
    • In patients homozygous for F508del mutation
    • Is promising but still inadequate
      • Decreased function or reduced PM stability of the pharmacologically-rescued F508del-CFTR (Van Goor et al., 2011, 2014; Clancy et al., 2012; Molinski et al., 2012)
  • LMTK2 knockdown in immortalized HBE cells
    • Did not increase the steady-state abundance of partially glycosylated F508del-CFTR band B
      • It did not rescue F508del-CFTR in the absence of VX-809


Lumacaftor


Methylxantiny

  • Inhibitory fosfodiesterázy
  • Působí bronchodilatačně a mírně protizánětlivě
  • Působí diureticky
  • Zvyšují srdeční výdej a mají mírný psychostimulační účinek

Retardované

Theofylin

  • V kombinované léčbě persistujícího astmatu zejména u pacientů s nočními příznaky
  • Pro své stimulační účinky na dechové centrum je theofylin používaný u pacientů s CHOPN
    • S opatrností by se měl podávat u pacientů s ICHS, srdeční nedostatečností, hypertenzí a hypertyreózou.

Neretardované methylxantiny

Aminophyllin

  • Zřídka podávány v akutních stavech nejčastěji v injekční formě
  • Bronchodilatační účinek je slabší oproti inhalačním beta2-mimetikům
  • Mohou však být prospěšné pro řízené dýchání a pro udržování odpovědi mezi dávkami RABA.

MgATP + Catalytic subunit of PKA

Milrinone and amrinone

  • Milrinone plus isoproterenol
    • In the transformed nasal polyp cells (CF-T43) homozygous for del F508-CFTR
    • Cl efflux could be stimulated [17]
  • Forskolin and milrinone to hyperpolarize the nasal epithelium
    • Indicative of a Cl- secretory response in ?F508 CFTR-expressing mice
  • DelF508 CFTR-expressing cells
    • Both an adenylyl cyclase agonist and milrinone were required to cause a response in vivo [17]
  • Forskolin and milrinone
    • Was ineffective in altering the nasal potential difference in the CFTR (-/-) mice [17]


Non-selective treatments

  • Low temperature (25–27°C)
  • Glycerol
  • 4-phenylbutyrate
  • Required long term exposure
    • To show modest effects on DF508-CFTR trafficking
  • [R L Dormer, C M Harris, Z Clark, M M C Pereira, I J M Doull, C Norez, F Becq, M A McPherson]

ORKAMBITM

Osthole

  • Del.F508-CFTR including [19]
  • Osthole showed the highest affinity with K(d) values pod 50 nmol/L as determined by Ussing chamber short-circuit current assay.
  • Stimulation of rat colonic mucosal secretion by osthole
    • Reached maximal activation of colonic Cl(-) secretion at 5 micro mol/L
  • Stimulation of mouse tracheal mucosal secretion
    • Fluid secretion rate of tracheal single submucosal gland stimulated by osthole at 10 micromol/L was three-fold more rapid than that in negative control.
  • Suggested the therapeutic potential of natural coumarin compounds for the treatment of CF and other CFTR-related diseases.
  • www.researchgate.net/publication/51710531_Stimulation_of_Airway_and_Intestinal_Mucosal_Secretion_by_Natural_Coumarin_CFTR_Activators


P2XRs extracellular ATP-gated, Ca2+-permeable, non-selective cation channels

  • Stimulation of airway epithelial P2XRs
    • Leads to sustained Ca2+ entry from extracellular stores
    • Dependent on extracellular pH and the presence or absence of extracellular calcium, sodium, and zinc
      • Degree of alkaline pH potentiation of zinc and ATP stimulation
  • Lack of desensitization or inactivation of the receptor or channel properties was novel
  • Combination of zinc and ATP
    • Could induce a prolonged Ca2+ signal
      • May rescue impaired Cl- secretion in CF airway epithelium in a sustained and reversible manner
  • www.jbc.org/article/S0021-9258(17)47733-7/fulltext

P2Y nucleotide receptors stimulation of G protein-coupled

  • Increases [Ca2+]i derived from intracellular stores
  • May affect both ion transport mechanisms
  • P2Y nucleotide receptors are currently a target for CF pharmacotherapy

PKC

  • Response of XCFTR to PKC stimulation is approximately sixfold that of PKA stimulation [19]
  • PKC activation does not cause insertion of new channels into the plasma membrane.[19]

Pentoxifylline

  • Pentoxifylline is a methyl-xanthine derivative
  • Inhibits the phosphodiesterase IV (PDE IV) = mohlo by to stimulovat CFTR (a zlepšit učinek KI ?)
  • Could have antiviral affects
  • Probable benefits of Pentoxifylline against COVID-19 pathogenesis
  • Many confirmatory data on proper efficacy of pentoxifylline on controlling COVID-19 and its consequences
  • Bronchodilator and respiratory supportive effects and protective roles in organ failures
  • Better circulation-oxygenation properties
  • Low price and safety
  • Make it a promising drug to be considered for COVID-19 treatment, especially as an adjuvant therapy in combination with other drugs
  • pubmed.ncbi.nlm.nih.gov/32473070/
  • Has previously been shown to inhibit several viral infections
  • Increases the levels of cyclic adenosine monophosphate
    • Activates protein kinase
      • Reduction in the synthesis of pro-inflammatory cytokines and immune cell migration
  • www.karger.com/Article/Abstract/512234

Phenylglycine PG-01


Phenylimidazothiazole drugs

  • Antihelminthic:
    • (-)-p-bromotetramisole
    • Levamisole [17]
  • Stimulated CFTR channel activity in CHO cells [17]
  • Alkaline phosphatase inactivates CFTR
    • Inhibition of a tonically active alkaline phosphatase accounted for stimulation
  • Stereospecific inhibition of alkaline phosphatase by phenylimidazothiazoles
    • 1.2 mcM for (-)-p-bromotetramisole
    • 11 mcM for levamisol [17]
  • Concentrations approaching 1 mM
    • Inhibited adenylyl cyclase [17]
  • Enhanced secretion may explain the ability of levamisole
    • To reduce the frequency, severity, and duration of upper respiratory tract infections in children [17]

Phosphatase inhibitors

  • Compounds that increase the activity of CFTR [16]
  • Inhibition of PP1, PP2A, or both results in the stabilization of an activated form of CFTR [17]

Nonspecific phosphatase inhibitors

  • F-
  • Metavanadate
  • Increased channel Po in this preparation
  • Inhibited the effects of exogenously applied alkaline phosphatase [17]
  • Vanadate
    • Had little or no observable effect on CFTR channel activity in excised membrane patches [17]

Type 1 phosphatases (PP1)

  • Preferentially dephosphorylate the beta-subunit of phosphorylase kinase
  • Sensitive to both inhibitory proteins
  • Okadaic acid, calyculin A, and microcystin
    • Selectivity for PP1 and PP2A [17]

Type 2 phosphatases (PP2)

  • Dephosphorylate the alpha-subunit
  • Sensitive to neither inhibitor [17]

PP2A

  • Requires no divalent cations [17]
  • Okadaic acid, calyculin A, and microcystin
    • Selectivity for PP1 and PP2A [17]

PP2B - also known as calcineurin

  • Are regulated by Ca2+ and calmodulin
  • Little effect of Ca2+ removal was noted
    • Indicating that PP2B did not modulate CFTR gating in excised membrane patches [17]
  • F- and phenothiazines
    • Selectively inhibit PP2B [17]
  • FK506 and cyclosporin
    • High-affinity inhibitors of PP2B [17]
    • After their interactions with binding proteins FKBP12 and cyclophilin [17]
    • FK506 did not inhibit cAMP-stimulated Cl- currents in airway or T84 colonic epithelial cell monolayers. [17]

PP2C

  • Require Mg2+ [17]
  • Orthovanadate selectively inhibits PP2C [17]
  • Removal of Mg2+
    • Alter CFTR gating in excised membrane patches [17]
    • Associated with significant reductions in the nucleotide-dependent opening rate of CFTR and in its closing rate [17]

Phosphodiesterase inhibitors

  • Compounds that increase the activity of CFTR [16]

Phosphodiesterase-type 5 (PDE5) inhibitors

  • (Dormer et al, 2005) sildenafil

Tyrosine phosphorylation

  • Contributes to muscarinic stimulation of CFTR [20]
  • 50% of CFTR’s response to carbachol persisted when PKA was inhibited by Rp-cAMPS [20]
  • Similar results wereobtained using a CFTR mutant that is unresponsive to forskolin [20]
  • Src Inhibitor-1 (Inh-1) abolished the PKA-independent component of muscarinic stimulation (Billetet al.2013) [20]

Pilocarpine

  • Ca2+-mobilizing muscarinic agonist [20]
  • Stimulates robust fluid secretion
  • This response is reduced by 60% in CF glands
    • Even more striking when one considers that it is probably an underestimate
      • More if normalized per unit area of epithelium to correct for 2- to 3-fold hyper-trophy of CF glands
  • Why pilocarpine-induced secretion should be compromised in CF submucosal glands if it occurs via CaCCs
    • Consistent observation in
      • Glands from CF piglet trachea (Jooet al.2010)
      • Nasal turbinate (Choet al.2011)
      • Tracheal xenografts (Sunet al.2010) [20]

Polymethoxylated flavonoids, 3',4',5,5',6,7-hexamethoxyflavone (HMF)

5-hydroxy-6,7,3',4'-tetramethoxyflavone (HTF)

  • Could potentiate CFTR chloride channel activities
  • Cell-based fluorescence assay and the short circuit Ussing chamber assay
    • Both HMF and HTF could dose-dependently potentiate CFTR Cl(-) channel activities in rapid and reversible ways
    • Activations could be reversed by the CFTR blocker CFTRinh-172
    • HMF showed the highest affinity (EC50 = 2 µmol/L) to CFTR protein among the flavonoid CFTR activators identified so far
    • Activation of CFTR by HMF or HTF was forskolin (FSK) dependent
    • Showed additive effect with FSK and 3-Isobutyl-1-methylx (IBMX) in the activation of CFTR
    • no additive effect with genistein (GEN)
  • HMF and HTF could stimulate transepithelial Cl(-) secretion in rat colonic mucosa and enhance fluid secretion in mouse trachea submucosal glands.
    • Through both elevation of cAMP level and binding to CFTR protein pathways [6]

Potentiators

  • Stimulate channel gating of mutant CFTR
  • Vx-770 (Van Goor et al., 2009)

Praeruptorin A

  • ?F508-CFTR including [19]

Protein kinases and phosphatases

  • Via phosphorylation and dephosphorylation
  • CFTR delivery to and retention in the plasma membrane

Psoralens

  • Compounds that increase the activity of CFTR [16]

Quercetin

  • Found to inhibit
    • PKC
    • Casein kinase
    • Phosphorylase kinase
    • Tyrosine kinase activity
    • Concentrations below 100 mM
      • Most “potent” kinase inhibitors available [17]

Quercetin and kaempferol

  • Stimulated Cl- secretion across T84 cell monolayers
    • Quercetin-induced stimulation was potentiated by carbachol, but not VIP
  • Quercetin produced a phosphoprotein map like that observed in the presence of cAMP
    • Effects were likely mediated by a PKA-dependent mechanism
    • CAMP production was unchanged [17]

Rab 4

  • Zvyšuje začlenění endozomu s CFTR do apikální membrány

Resveratrol (RES)

  • RES could induce intestinal chloride secretion in mouse jejunum
  • Stimulate cAMP-dependent Cl- secretion in T84, primary cultured murine nasal septal and human sinonasal epithelial cells
  • Cell-based fluorescent assays, transepithelial short-circuit current measurements and excised inside-out patch-clamp analysis
    • RES dose-dependently potentiate CFTR Cl- channel activities
    • Reversed by CFTR inhibitors CFTR(inh)-172 and GlyH101
    • Stimulated by RES with half maximal concentration -80 microM
    • Intracellular cAMP content was not elevated by RES in FRT cells
    • RES significantly increased the chloride currents of CFTR
    • RES stimulated the transmucosal chloride current of rat colon by short-circuit current assay
    • RES may represent a novel class of therapeutic lead compounds in treating CFTR-related diseases including CF and habitual constipation. [11]

May act by modifying keratin 18 (K18) network (Hamdaoui et al, 2011; Lipecka et al, 2006)

  • K18 heterodimerizes with keratin 8 (K8)
  • Role in the trafficking of CFTR/ DF508-CFTR (Colas et al, 2012; Duan et al, 2012).
    • Decrease in K8 expression leads to functional correction of DF508-CFTR (Colas et al, 2012).
    • [EMBO Mol Med (2013) 5, 1484–1501, 2013 The Authors. Published by John Wiley and Sons, Ltd on behalf of EMBO, Research Article Norbert Odolczyk et al.]

Roscovitine

  • Tested in clinical studies as therapeutic option for a plethora of diseases
  • Partial corrector of the F508del CFTR protein
  • Recruits TRPC6 translocation to phagosomal membranes
    • Able to restore microbicidal function in alveolar macrophages compromised by CF
  • Other functions of this drug on neutrophils, epithelial cells, eosinophils, and lymphocytes
    • May also be beneficial to restore activity in CF affected cells
  • www.mdpi.com/1424-8247/12/1/23/htm

S-Nitrosoglutathione

  • Endogenous bronchodilator and signaling molecule
  • Enhances expression, maturation, and function of both wt and deltaF508 CFTR in epithelial cells
  • Present endogenously on the apical side of airway epithelium
  • Increases ciliary beat frequency
    • Improving mucociliary clearance
  • Nitric oxide synthases (NOSs) are involved in conversion of L-arginine into nitric oxide (NO)
  • NO, in turn, is involved in production of S-nitrosothiols
    • Including S-Nitrosoglutathione
  • NO may react directly with thiyl radicals or with thiols to form S-nitrosothiols or S-nitrosothiol radicals
  • Three isoforms of NOS:
    • Neuronal NOS (nNOS),
    • Endothelial NOS (eNOS)
    • Inducible NOS (iNOS) -by inflammatory stimuli
      • Synthesizes relatively large quantities of NO
  • All three isoforms are expressed in human airways
  • Polymorphisms of constitutive NOS (nNOS and eNOS) and reduced iNOS expression
    • Contributes to decreased NO production along with bacterial consumption
  • S-nitrosylation
    • Can functionally regulate the general activities of Heat shock protein 90kDa alpha (HSP90 alpha)
    • Provide a feedback mechanism for limiting eNOS activation
  • S-Nitrosoglutathione
    • Covalently modifies a susceptible cysteine residue in the HSP90 alpha domain that interacts with eNOS
    • Direct S-nitrosylation can increase the activity of each of the major forms of nitric oxide synthases (nNOS, eNOS and iNOS)
  • Ceruloplasmin
    • Also may catalyze the synthesis of S-Nitrosoglutathione
  • S-Nitrosoglutathione
    • Can be catabolized by a number of enzymes
      • Cu/Zn superoxide dismutase (SOD1),
      • Gamma glutamyl transpeptidase (Gamma GT)
      • Thioredoxin reductases (TXNRD1, TXNRD2 and TXNRD3)
      • Glutathione-dependent formaldehyde dehydrogenase (ADHX (GSNOR)

Gamma GT can be involved in CFTR activation

  • S-Nitrosocysteinylglycine
    • Product of S-Nitrosoglutathione cleavage by Gamma GT, can increase DeltaF508 CFTR maturation
  • SOD1, TXNRD1, TXNRD2 and TXNRD3 catabolize S-Nitrosoglutathione
    • To form free NO radicals
  • Free NO can spontaneously react with Superoxide anion radical (O(2)(-)) to produce Peroxynitrite (ONOO(-))
    • SOD1
      • Catalyzes the dismutation of O(2)(-)
      • Can outcompete the peroxynitrite reaction
    • Cells may contain sufficient SOD1 to prevent inactivation of NO by O(2)(-)
  • S-Nitrosoglutathione at low micromolar concentrations
    • Increases the DeltaF508 and wild-type CFTR expression and maturation
    • Mainly acts independently of the classic NO radical/cyclic GMP pathway
  • Effect of S-Nitrosoglutathione at 1-10 microM concentration
    • Increasing transcription factors SP1 and SP3 expression and their DNA-binding capacity
    • Post-translational
  • SP1
    • Additional mechanism for enhanced DNA-binding involves cysteine S-nitrosylation in the SP1 zinc finger-binding domain
  • Post-translational effect of S-Nitrosoglutathione
    • Increased expression and covalent modification - namely S-nitrosylation - of proteins involved in CFTR folding, and stabilization resulted in an increased CFTR maturation
  • ER-associated pathways of CFTR folding are affected by chaperones and co-chaperones such as cytosolic Heat shock proteins 70 and 90kDa (HSP70 and HSP90), DnaJ homolog subfamily B member (Hdj-1)
  • HSP90 (HSP90 alpha and HSP90 beta) and Heat shock 70kDa protein 8 (HSC70)
    • S-nitrosylated by S-Nitrosoglutathione
      • Followed by CFTR folding and stabilization
  • S-Nitrosoglutathione also increases expression of DnaJ homolog, subfamily C, member 5 (Csp)
    • To enhance the association between Csp and CFTR in the ER and Golgi
  • S-Nitrosoglutathione increases Csp expression (primarily post-transcriptionally) leading to increase in CFTR folding and maturation
    • Csp initiates activation of HSC70 ATPase activity, which leads to CFTR degradation
      • Degradation is inhibited in the presence of S-Nitrosoglutathione, allowing increased Csp to continue stabilization of CFTR
  • HSC70 has a single critical cysteine residue in its ATP binding domain
    • S-nitrosylation of this cysteine allows Csp to augment CFTR folding without leading to CFTR degradation

S-Nitrosoglutathione at nitrosative stress levels (100 microM)

  • Inhibits SP3 binding
  • Augments competitive binding of SP1
  • Inhibits CFTR transcription
    • cftr-activation-s-nitrosoglutathione-normal-cf">www.bio-rad.com/en-cz/prime-pcr-assays/pathway/mechanisms-cftr-activation-s-nitrosoglutathione-normal-cf

SARS-CoV-2

  • CFTR and ENaC proposed as theoretical cleavage sites for the coronavirus proteinase 3CLpro enzyme
  • Transmembrane protease serine 2 (TMPRSS2)
    • Facilitate viral entry into the target host cell
    • Reduces ENaC activity in airway epithelium
  • www.nature.com/articles/s41435-020-0103-y

Scoparone

  • ?F508-CFTR including [19]

Serine proteases

Sildenafil (Viagra)

  • Corrects DF508-CFTR location in nasal epithelial cells from patients with cystic fibrosis
  • In most untreated CF cells DF508-CFTR was mislocalised within the cell at a site close to the nucleus
  • Exposure of cells to sildenafil resulted in
    • Recruitment of DF508-CFTR to the apical membrane
    • Appearance of chloride transport activity
  • Sildenafil also increased DF508-CFTR trafficking in cells from individuals with
    • CF with a single copy DF508 (DF508/4016ins)
    • With a newly described CF trafficking mutation (R1283M)
  • [R L Dormer, C M Harris, Z Clark, M M C Pereira, I J M Doull, C Norez, F Becq, M A McPherson]

Sphingomyelin complexed to CFTR

  • Promotes gating of the channel
  • CFTR channel reconstituted in a (nonsphingomyelin) phospholipid bilayer
    • Exhibits relatively high P
      • Retains its native sphingomyelin
      • And/or that other phospholipids with a suitable head group can also promote CFTR gating
  • www.pnas.org/content/104/15/6448

Ivacaftor plus tezacaftor (Symdeko™)


TNF alfa

  • 10 min exposure to TNF-alpha (0.5-50ng/ml) of F508del-CFTR-transfected HeLa cells and human bronchial cells expressing F508del-CFTR in primary culture
    • Leads to the maturation of F508del-CFTR
    • Induces CFTR chloride currents
    • Sustained, in HBE cells, for at least 24 h
    • Involves a protein kinase C (PKC) signaling pathway
    • TNF? behaving as a corrector molecule
  • Novel and unexpected action of TNFalpha
  • Calu-3 cells derived from a pulmonary adenocarcinoma - TNFalpha activated CFTR gene expression at the transcriptional level
  • Colon adenocarcinoma-derived cell line (T84) - TNFalpha - reduced CFTR expression
  • Residual activity of CFTR in the nasal epithelium exists in patients with a mild phenotype
    • Suggesting that inflammatory status may be correlated with residual CFTR function
  • IL-1beta has recently been reported to stimulate the expression of CFTR in T84 colon carcinoma cells
  • IL-1beta, another pro-inflammatory cytokine, did not affect I CFTR
  • Chronic treatment of intestinal cells by TNFalpha (>24h)
  • Ha zdá se, že u každého člověka to může být jiné dle toho, jakou má mutaci nebo nemá




Tangeretin from Pericarpium Citri Reticulatae Viride

  • CFTR is a cAMP-activated chloride channel
  • Expressed in the apical membrane of serous epithelial cells
  • CFTR activator
    • Identifikován via high-throughput screening based on FRT cell-based fluorescence assay
  • Fluorescence quenching tests showed that
    • Tangeretin dose- and time-dependently activated CFTR chloride channel
    • Activity had rapid and reversible characteristics
    • Activation effect could be completely reversed by the CFTR specific blocker CFTRinh-172 [5]
  • Primary mechanism studies indicated
    • FSK dependent
    • Additional effect with FSK and IBMX
      • = tangeretin activates CFTR by direct interacting with the protein [5]
  • Ex-vivo tests
    • Tangeretin could accelerate the speed of the submucosal gland fluid secretion [5]
  • Short-circuit current measurement
    • Tangeretin activated rat colonic mucosa chloride current
    • CFTR Cl(-) channel is a molecular target of natural compound tangeretin [5]
  • May have potential use for the treatment of CFTR-related diseases like
    • Cystic fibrosis,
    • Bronchiectasis
    • Habitual constipation [5]

Trimethylamine oxide

Tyrphostin B42

  • Structurally unrelated tyrosine kinase inhibitor
  • Had similar effects to those of genistein [17]
  • Genistein and tyrphostin 47
    • Stimulated short-circuit current across T84 cell monolayers with an EC50 of 12.5 mcM [17]

Vasoactive intestinal peptide

  • (VIP) is a 28–amino acid neuropeptide released by
    • Nonadrenergic,
    • Noncholinergic neurons that innervate the airways (Wine, 2007)
    • By immune cells (Martinez et al., 1999)
  • VIP binds to the G protein–coupled receptors VPAC1 and VPAC2 (vasoactive intestinal peptide receptor types 1 and 2)
    • On airway epithelial cells (Groneberg et al., 2001; Dérand et al., 2004; Miotto et al., 2004)
  • Leading to activation of PKA and PKC signaling
    • Stimulation of CFTR-mediated secretion (Schwartz et al., 1974; Dharmsathaphorn et al., 1985) [22]



VX-809

  • Small but similar to that observed in previous studies (Van Goor et al., 2011; Cihil et al., 2013; Holleran et al., 2013)
  • Attenuating LMTK2 phosphorylation of F508del-CFTR
    • May play a role together with CFTR correctors and potentiators to maximally rescue F508del-CFTR function in CF patients
  • www.frontiersin.org/articles/10.3389/fchem.2016.00001/full

Vitamin C (l-ascorbate)

  • Present in the respiratory lining fluid of human lungs
  • Local deficits occur during oxidative stress
  • Unique function of vitamin C on the cystic fibrosis (CF) transmembrane conductance regulator (CFTR)
  • Vitamin C (100 mcroM) induced the openings of CFTR Cl channels
    • By increasing its average open probability from 0 to 0.21 ± 0.08
    • Without a detectable increase in intracellular cAMP levels
  • Exposure of the apical airway surface to vitamin C stimulated the transepithelial Cl secretion to 68% of forskolin-stimulated currents
  • The average half-maximal stimulatory constant was 36.5 ± 2.9 mcM
    • Corresponds to physiological concentrations
  • When vitamin C was instilled into the nasal epithelium of human subjects
    • It effectively activated Cl transport in vivo
  • In CF epithelia, previous treatment of the underlying trafficking defect with trimethylamine oxide or expression of WT CFTR
    • Restored the
      • Activation of Cl transport by vitamin C
      • Sodium dependency and phloretin sensitivity
      • Expression of transcripts for sodium-dependent vitamin C transporter (SVCT)-1 and SVCT2
  • We conclude that cellular vitamin C is a biological regulator of CFTR-mediated Cl secretion in epithelia
  • Pool of vitamin C in the respiratory tract represents a potential nutraceutical and pharmaceutical target for the complementary treatment of sticky airway secretions by enhancing epithelial fluid secretion.

Xanthines

  • Stimulate CFTR by modulating phosphatase activity [17]

Extracellular Zinc and ATP

  • Restore Chloride Secretion across Cystic Fibrosis Airway Epithelia by Triggering Calcium Entry
  • Zinc and ATP induced a sustained, reversible, and reproducible increase in cytosolic Ca2+ in CF and non-CF cells
    • Activation of P2X purinergic receptor channels
      • Mediated Ca2+ entry stimulated sustained Cl- and HCO-3 secretion in CF and non-CF epithelial monolayers
  • www.jbc.org/article/S0021-9258(17)47733-7/fulltext

Zinc supplementation

  • Can improve CF phenotypes (respiratory function and the incidence of infection) (Van Biervliet et al., 2008, Abdulhamid et al., 2008)
  • Zinc dysregulation may also be a possible pathophysiological hallmark of CF.
  • Zinc deficiency via the unique splicing switch of ZIP2 is crucial for CF lung pathology
    • Especially with respect to the induction of MUC5AC
      • A major secreted mucin that is highly expressed in CF airway
        • Interplay of CFTR, ENaC, and ZIP2 transporters.
  • www.sciencedirect.com/science/article/pii/S2352396417305066

Low levels of acetylcholine

  • ACh, a Ca2+-elevating agonist
  • Hyperpolarize the intracellular potential
    • By activating basolateral K+ channels
    • Thereby maintaining the driving force for Cl- secretion
  • This leads to synergistic effects on secretion
  • Failure to respond to either or both agents may promote bacterial colonization [26]

Butyrát

  • Of cell cultures with sodium butyrate or sodium 4-phenylbutyrate at non-toxic concentrations
    • Increases normal CFTR processing
    • Leads to cAMP-mediated chloride conduction at the cell surface
  • Butyrate has a short half-life in vivo
    • Must be administered by intravenous drip
  • Induce fetal hemoglobin
  • Induce differentiation in a variety of solid tumors

Phenylbutyrate

  • Is an oral pro-drug of phenylacetate
    • Also capable of inducing fetal hemoglobin
    • Acting as an adjunct chemotherapeutic agent
    • Inducing CFTR expression and trafficking
  • It does not appear to affect CFTR single channel properties
  • phenylbutyrate-mediated reduction in Hsc70
    • To explain the shift of mutant CFTR onto the trafficking pathway
  • Both DSG and the butyrates may regulate interactions of CFTR with ER chaperones
  • Randomized, double-blind, placebo-controlled trial in CF patients homozygous for del F508
    • One week of phenylbutyrate at 20 gm/day
      • Associated with induction of chloride transport based on nasal potential difference measurements
    • no safety issues arose in the CF trial
      • Widespread disruption of the quality control mechanism for folding or degrading proteins was not detectable

Buphenyl (4-phenylbutyrate)

  • Newly approved drug for the rare inherited urea cycle disorders
    • Provide an alternative vehicle for waste nitrogen excretion by conjugation with glutamine
  • Under study in the hemoglobinopathies, adrenoleukodystrophy, cancer trials
  • Dose escalation and pharmacokinetic studies are continuing in adults with CF.
  • www.ncbi.nlm.nih.gov/pmc/articles/PMC408112/


cAMP, Ca2+ or cGMP

  • Inhibit electroneutral Na+ absorption and activate Cl- secretion in intestine [18]

Stimulation of the cAMP signal transduction cascade

  • Stimuluje začlenění endozomů s CFTR do membrány (podobně jako GLUT4)
  • Channel activation can arise [16]
  • CFTR is primarily activated by:
    • PKA-dependent phosphorylation of its unique regulatory (R) domain
    • ATP binding/hydrolysis at 2 nucleotide binding domains, which form a heterodimer to gate channel activity
  • CFTR function, therefore, is dependent on local: 3',5'-cyclic adenosine monophosphate (cAMP) accumulation
    • Highly regulated by spatially restricted proteins in the region of the CFTR
  • Acute cAMP elevation is a necessary step in physiologic CFTR activation
    • Loss of regulation leads to pathology such as
      • Cholera-induced diarrheal disease - cholera toxin bypasses the cAMP regulatory network to chronically raise cAMP - life-threatening fluid losses
  • CAMP is produced by ACs
    • A group of enzymes with 9 identified transmembrane isoforms
      • 2 hydrophobic domains
      • 6 transmembrane spans
      • 2 cytoplasmic domains - catalytic activity
  • All 9 transmembrane AC isoforms are activated by the GSalpha subunit of the GP
    • All but AC9 are directly activated by the diterpene forskolin
  • insight.jci.org/articles/view/93029
  • Acute cAMP elevation activates CFTR in vitro and in vivo
  • 95% of CF patients are treated daily with beta 2AR-agonists (which raise cAMP in airway epithelia)
    • Despite a recommendation of “insufficient evidence to support chronic use” from the CF Foundation
  • Short-acting ß2AR-agonists, such as albuterol
    • Frequently used as a part of daily airway clearance regimens
  • Long-acting ß2AR-agonists (LABAs — such as formoterol)
    • Also commonly prescribed
      • Often in combination with inhaled corticosteroids to control asthma symptoms
  • insight.jci.org/articles/view/93029
  • Localized cAMP concentration in the vicinity of the channel
  • Complex arrangement fine tunes the spatio-temporal regulation of CFTR phosphorylation-dephosphorylation events
  • Influenced by multidrug resistance-associated proteins, e.g. the
    • Multidrug resistance-associated protein 4 (MRP4) activity
    • ABC transporter that can extrude cytosolic cAMP
  • www.nature.com/articles/s41598-019-48971-y
  • Del F508 CFTR is capable of transporting Cl– in response to cAMP agonists
  • In vivo and ex vivo studies show that respiratory and intestinal tissues from ?F508 CFTR homozygous
    • Are competent to respond to agonists of the cAMP-dependent Cl– secretory pathway
  • J. Clin. Invest. 108:1705–1715 (2001). DOI:10.1172/JCI200112108.

cAMP via activation of adenylate cyclase by a G-protein

  • Glucagon
  • Epinephrine
  • Beta-adrenergic agonist
    • Isoproterenol
    • Acetylcholine
    • The vasoactive intestine peptide (VIP)
    • Adenosine
  • Intracellular cAMP obtained by hormonal stimulation
    • Maximal activity of CFTR, might be well below the levels obtained by
      • Forskolin or permeant cAMP analogues.
  • Forskolin
    • (0.5–10 mcM) leads to an intracellular increase of cAMP sufficient to activate the PKA
    • High concentrations of forskolin (N20 µM) may produce unspecific responses,
      • Triggers a feedback mechanism
        • Initial fast increase followed

Inhibition of inactivating enzymes (phosphodiesterases, phosphatases)

  • Channel activation can arise [16]

PKA-mediated phosphorylation of the regulatory (R) domain

Secretagogues Calu-3 cell monolayers

  • Generate robust increases in short-circuit current (Isc)
    • Due to elevation of cAMP
      • Isoproterenol
      • Forskolin
    • calcium
      • Bradykinin
      • histamine
      • Trypsin
      • Methacholine
      • Ca2+ ionophores (Shen et al., 1994). [27]

Stimulace CFTR transportéru

  • Inhibitor S-nitrosoglutathion reduktázy (GSNOR inhibitor)
    • Vyšší hladiny S-nitrosoglutathionu (vznikají při vyšších hladinách glutathionu)
  • Phloxin B (PB)
  • S-nitrosoglutathion diethyl ester (GNODE)
    • Stimulace maturace a stabilizace v membráně
  • S-nitro-N-acetylcystein (SNOAC)
    • Stimulace maturace a stabilizace v membráně

The tricyclic nucleotides, Uridine triphosphate and adenosine triphosphate

  • Regulate ion transport through P2Y2 purinergic receptors

Uridine triphosphate aerosol

  • Alone or in combination with amiloride
  • Increases transepithelial potential difference and the clearance of inhaled radioaerosol

P2Y2 purinergic receptor agonists


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Poslední aktualizace: 27. 2. 2021 23:04:50