Dual Therapeutic Utility of Proteasome Modulating Agents for Pharmaco-gene Therapy of the Cystic Fibrosis Airway

Zhang, Liang; Karp, Phil; Gerard, Christopher J.; Pastor, Eric; Laux, Douglas; Munson, Keith; Yan, Ziying; Liu, Xiaoming; Godwin, Simon; Thomas, Christie P.; Zabner, Joseph; Shi, Huidong; Caldwell, Charles W.; Peluso, Richard W.; Carter, Barrie; Engelhardt, John F.

doi:10.1016/j.ymthe.2004.08.009

Cited by 46 publications

(51 citation statements)

References 41 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Indeed, our (and other groups) identification of HspA4, HspBP1, and other chaperones as proteins when overexpressed correct F508del-CFTR cell surface expression and function led us to test the ability of Velcade (a proteasome inhibitor that activates the cellular stress response, including stimulation of chaperones) to rescue F508del-CFTR. Our work here shows that Velcade can indeed correct F508del-CFTR function, in accordance with a recent report by Zeitlin and colleagues (43) and with a report that demonstrates rescue of mutant CFTR with the proteasome inhibitor LLnL (44). Since Velcade is already used clinically for the treatment of multiple myeloma, it may be possible to test its efficacy for treating CF.…”

Section: Discussionsupporting

confidence: 92%

High-content Functional Screen to Identify Proteins that Correct F508del-CFTR Function

Trzcińska-Daneluti¹,

Ly²,

Huynh

et al. 2009

Molecular & Cellular Proteomics

View full text Add to dashboard Cite

Cystic Fibrosis is caused by mutations in CFTR, with a deletion of a phenylalanine at position 508 (F508del-CFTR) representing the most common mutation. The F508del-CFTR protein exhibits a trafficking defect and is retained in the endoplasmic reticulum. Here we describe the development of a high-content screen based on a functional assay to identify proteins that correct the F508del-CFTR defect. Using a HEK293 MSR GripTite cell line that stably expresses F508del-CFTR, we individually co-expressed ϳ450 unique proteins fused to the Cl ؊ -sensitive YFP(H148Q/I152L) mutant. We then tested correction of F508del-CFTR function by the CI ؊ /l ؊ exchange method following stimulation with forskolin/IBMX/genistein, using quantitative recordings in multiple individual cells with a high-content (high-throughput) Cellomics KSR imaging system. Using this approach, we identified several known and novel proteins that corrected F508del-CFTR function, including STAT1, Endothelin 1, HspA4, SAPK substrate protein 1, AP2M1, LGALS3/galectin-3, Trkfused gene, Caveolin 2, PAP/REG3␣, and others. The ability of these correctors to rescue F508del-CFTR trafficking was then validated by demonstrating their enhancement of maturation (appearance of band C) and by cell surface expression of F508del-CFTR bearing HA tag at the ectodomain using confocal microscopy and flow cytometry. These data demonstrate the utility of highcontent analyses for identifying proteins that correct mutant CFTR and discover new proteins that stimulate this correction. This assay can also be utilized for RNAi screens to identify inhibitory proteins that block

show abstract

Section: Discussionsupporting

confidence: 92%

High-content Functional Screen to Identify Proteins that Correct F508del-CFTR Function

Trzcińska-Daneluti¹,

Ly²,

Huynh

et al. 2009

Molecular & Cellular Proteomics

View full text Add to dashboard Cite

show abstract

“…Four to six weeks after ALI was established, transepithelial short circuit currents (I sc ) were measured using an epithelial voltage clamp (Model EC-825) and a self-contained Ussing chamber system (both purchased from Warner Instruments, Inc., Hamden, CT) as previously described (35,36). Throughout the experiment the chamber was kept at 37ЊC, and the chamber solution was aerated.…”

Section: Short Circuit Current Measurementsmentioning

confidence: 99%

Bioelectric Properties of Chloride Channels in Human, Pig, Ferret, and Mouse Airway Epithelia

Liu

Luo

Zhang

et al. 2007

Am J Respir Cell Mol Biol

Self Cite

View full text Add to dashboard Cite

The development of effective therapies for cystic fibrosis (CF) requires animal models that can appropriately reproduce the human disease phenotype. CF mouse models have demonstrated cAMPinducible, non-CF transmembrane conductance regulator (non-CFTR) chloride transport in conducting airway epithelia, and this property is thought to be responsible for the lack of a spontaneous CF-like phenotype in the lung. Thus, an understanding of species diversity in airway epithelial electrolyte transport and CFTR function is critical to developing better models for CF. Two species currently being used in attempts to develop better animal models of CF include the pig and ferret. In the study reported here, we sought to comparatively characterize the bioelectric properties of in vitro polarized airway epithelia-from human, mouse, pig and ferretgrown at the air-liquid interface (ALI). Bioelectric properties analyzed include amiloride-sensitive Na ؉ transport, 4,4-diisothiocyanato-stilbene-2,2-disulfonic acid (DIDS)-sensitive Cl ؊ transport, and cAMP-sensitive Cl ؊ transport. In addition, as an index for CFTR functional conservation, we evaluated the ability of four CFTR inhibitors, including glibenclamide, 5-nitro-2-(3-phenylpropyl-amino)-benzoic acid, CFTR inh -172, and CFTR inh -GlyH101, to block cAMPmediated Cl ؊ transport. Compared with human epithelia, pig epithelia demonstrated enhanced amiloride-sensitive Na ϩ transport. In contrast, ferret epithelia exhibited significantly reduced DIDS-sensitive Cl ؊ transport. Interestingly, although the four CFTR inhibitors effectively blocked cAMP-mediated Cl ؊ secretion in human airway epithelia, each species tested demonstrated unique differences in its responsiveness to these inhibitors. These findings suggest the existence of substantial species-specific differences at the level of the biology of airway epithelial electrolyte transport, and potentially also in terms of CFTR structure/function.

show abstract

“…A number of strategies have been explored to circumvent this limitation. Some studies have used short endogenous AAV sequences as a promoter (with or without proteosomal inhibitors to increase expression) with variable results in differentiated airway epithelia (15,18,19). Other studies have produced AAV vectors with a CFTR cDNA that deletes the N terminus (15), and other deletions have been proposed for AAV vectors (16,18).…”

mentioning

confidence: 99%

A shortened adeno-associated virus expression cassette for CFTR gene transfer to cystic fibrosis airway epithelia

Ostedgaard

Rokhlina

Karp

et al. 2005

Proc. Natl. Acad. Sci. U.S.A.

Self Cite

View full text Add to dashboard Cite

Adeno-associated viruses (AAVs) such as AAV5 that transduce airway epithelia from the apical surface are attractive vectors for gene transfer in cystic fibrosis (CF). However, their utility in CF has been limited because packaging of the insert becomes inefficient when its length exceeds Ϸ4,900 -5,000 bp. To partially circumvent this size constraint, we previously developed a CF transmembrane conductance regulator (CFTR) transgene that deleted a portion of the R domain (CFTR⌬R). In this study, we focused on shortening the other elements in the AAV expression cassette. We found that portions of the CMV immediate͞early (CMVie) enhancer͞promoter could be deleted without abolishing activity. We also tested various intervening sequences, poly(A) signals, and an intron to develop an expression cassette that meets the size restrictions imposed by AAV. We then packaged these shortened elements with the CFTR⌬R transgene into AAV5 and applied them to the apical surface of differentiated CF airway epithelia. Two to 4 weeks later, the AAV5 vectors partially corrected the CF Cl ؊ transport defect. These results demonstrate that a single AAV vector can complement the CF defect in differentiated airway epithelia and thereby further the development of effective CF gene transfer.

show abstract

Dual Therapeutic Utility of Proteasome Modulating Agents for Pharmaco-gene Therapy of the Cystic Fibrosis Airway

Cited by 46 publications

References 41 publications

High-content Functional Screen to Identify Proteins that Correct F508del-CFTR Function

High-content Functional Screen to Identify Proteins that Correct F508del-CFTR Function

Bioelectric Properties of Chloride Channels in Human, Pig, Ferret, and Mouse Airway Epithelia

A shortened adeno-associated virus expression cassette for CFTR gene transfer to cystic fibrosis airway epithelia

Contact Info

Product

Resources

About