ABCMdb

PMID: 18595696

Al-Nakkash L, Springsteel MF, Kurth MJ, Nantz MH
Activation of CFTR by UCCF-029 and genistein.
Bioorg Med Chem Lett. 2008 Jul 15;18(14):3874-7. Epub 2008 Jun 19., [PubMed]

Sentences

No. Mutations Sentence Comment

3 ABCC7 p.Lys1250Ala Using NIH3T3 cells stably transfected with K1250A-CFTR we find that CFTR channel open time is unaffected by UCCF-029 or genistein, supporting the hypothesis that these compounds stabilize the open state by inhibiting ATP hydrolysis at NBD2. Login to comment 56 ABCC7 p.Lys1250Ala ABCC7 p.Lys1250Ala To examine whether UCCF-029 acts to stabilize the channel open state, we determined its effect upon the K1250A-CFTR channel (a CFTR channel mutation that can stay open for minutes once opened).16 A representative recording of K1250A-CFTR in cell-attached patch is shown (Fig. Login to comment 62 ABCC7 p.Lys1250Ala In addition, we demonstrate that UCCF-029 is unable to further stimulate the K1250A-CFTR chloride current activated by a maximally effective concentration of forskolin. Login to comment 63 ABCC7 p.Lys1250Ala ABCC7 p.Lys1250Ala In addition, we demonstrate that UCCF-029 is unable to further stimulate the K1250A-CFTR chloride current activated by a maximally effective concentration of forskolin. Login to comment 64 ABCC7 p.Lys1250Ala We have previously shown similar results with genistein and benzimidazolone analogs.6 Our earlier data using relaxation analysis demonstrated that these compounds stabilize open state by inhibiting ATP hydrolysis at NBD2.6 K1250A-CFTR, a mutation of the Walker A lysine at NBD2 prolongs channel opening by eliminating ATP hydrolysis at NBD2.19,16 The original work describing UCCF-029 compared its effects to several other novel compounds generated using combinatorial libraries and with genistein,11 using the halide-sensitive yellow fluorescent protein on Wt-CFTR transfected Fischer rat thyroid (FRT) cells. Login to comment 65 ABCC7 p.Gly551Asp However, the difference in effectiveness of UCCF-029 in our hands versus theirs could be attributed to the following: (1) we used NIH3T3 cells transfected with DF508-CFTR, whereas they used FRT cells transfected with Wt-CFTR (interestingly in those same studies, UCCF-029 was not as effective an activator on another CFTR mutation G551D), (2) our experiments used single isolated cells, whereas they used monolayers of cells either grown in 96-well plates for the fluorescent assays or polarized monolayers for Ussing electrophysiological short circuit current measures. Login to comment 66 ABCC7 p.Gly551Asp However, the difference in effectiveness of UCCF-029 in our hands versus theirs could be attributed to the following: (1) we used NIH3T3 cells transfected with DF508-CFTR, whereas they used FRT cells transfected with Wt-CFTR (interestingly in those same studies, UCCF-029 was not as effective an activator on another CFTR mutation G551D), (2) our experiments used single isolated cells, whereas they used monolayers of cells either grown in 96-well plates for the fluorescent assays or polarized monolayers for Ussing electrophysiological short circuit current measures. Login to comment