Yes / Type II congenitalLong QT syndrome (LQT2) is due to genetic mutations in hERG channel. Genetic or
pharmacological factors could potentially affect hERG channel biogenesis and contributes to LQTS, for example,
disease mutations G601S and T473P result in hERG trafficking deficiency [1,2]. Various rescue strategies for hERG
dysfuction are being developed. Some correctors for CFTR channel have been reported to act indirectly on
proteostasis pathways to promote folding and correction on hERG trafficking deficiency [3]. In this study, we tested the
hypothesis that the CFTR corrector KM-11060 and the potentiator PG-01 may correct hERG mutation trafficking
diseases.
We use HEK293 cell line expressing a well-studied trafficking disease mutation G601S-hERG channel [4]. We treated
cells with CFTR potentiator PG-01and corrector KM-11060, which function through different cellular mechanisms, and
assessed whether correction occurred via immunoblotting. Whole cell proteins from HEK 293 cells expressing hERG
channels were used for analysis [5]. Proteins were separated on 8% SDS-polyacrylamide electrophoresis gels for 1
hour, transferred onto PVDF membrane, and blocked for 1 h with 5% nonfat milk. The blots were incubated with the
primary antibody (Santa Cruz Biotechnology) for 12-16 h at 4C temperature and then incubated with a donkey antigoat
horseradish peroxidase-conjugated secondary antibody( Santa Cruz Biotechnology). Actin expression was used
for loading controls. The blots were visualized using the ECL detection kit (Genshare).Results were deemed
significantly different from controls by a one-way ANOVA (p < 0.05).
Our results show that both KM-11060 (5, 10, 20uM) and PG-01(5, 15 uM) can correct G601S mutant alleles of hERG
protein trafficking (Fig 1, 2). KM-11060 (20uM) but not PG-01(15 uM) enhance protein expression of wild type hERG
channel (Fig 2). Further treatment on cells at low temperature with different drug concentration will be tested.
Functional studies are also needed to test whether the drugs can correct the function of hERG mutation channel.
These results could potentially provide novel insight into the correction mechanism of CFTR potentiator and also help
to develop new treatment for LQT2.
| Identifer | oai:union.ndltd.org:BRADFORD/oai:bradscholars.brad.ac.uk:10454/10856 |
| Date | January 2016 |
| Creators | Zhang, J., Shang, Lijun, Ma, A. |
| Source Sets | Bradford Scholars |
| Language | English |
| Detected Language | English |
| Type | Conference paper, Accepted manuscript |
| Rights | © 2016 The Authors. Published by the Physiological Society. Reproduced in accordance with the publisher's self-archiving policy., Unspecified |
| Relation | http://www.physoc.org/proceedings/abstract/Proc%20Physiol%20Soc%2037PCB120 |
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