Coexistence of hERG current block and disruption of protein trafficking in ketoconazole-induced long QT syndrome.
OBJECTIVE
Many drugs associated with acquired long QT syndrome (LQTS) directly block human ether-a-go-go-related gene (hERG) K(+) channels. Recently, disrupted trafficking of the hERG channel protein was proposed as a new mechanism underlying LQTS, but whether this defect coexists with the hERG current block remains unclear. This study investigated how ketoconazole, a direct hERG current inhibitor, affects the trafficking of hERG channel protein.


METHODS
Wild-type hERG and SCN5A/hNa(v) 1.5 Na(+) channels or the Y652A and F656C mutated forms of the hERG were stably expressed in HEK293 cells. The K(+) and Na(+) currents were recorded in these cells by using the whole-cell patch-clamp technique (23 degrees C). Protein trafficking of the hERG was evaluated by Western blot analysis and flow cytometry.


RESULTS
Ketoconazole directly blocked the hERG channel current and reduced the amount of hERG channel protein trafficked to the cell surface in a concentration-dependent manner. Current density of the hERG channels but not of the hNa(v) 1.5 channels was reduced after 48 h of incubation with ketoconazole, with preservation of the acute direct effect on hERG current. Mutations in drug-binding sites (F656C or Y652A) of the hERG channel significantly attenuated the hERG current blockade by ketoconazole, but did not affect the disruption of trafficking.


CONCLUSIONS
Our findings indicate that ketoconazole might cause acquired LQTS via a direct inhibition of current through the hERG channel and by disrupting hERG protein trafficking within therapeutic concentrations. These findings should be considered when evaluating new drugs.
Takemasa
H
H
Second Department of Internal Medicine, University of Occupational and Environmental Health Japan, Yahatanishi-ku, Kitakyushu, Japan.
Nagatomo
T
T
Abe
H
H
Kawakami
K
K
Igarashi
T
T
Tsurugi
T
T
Kabashima
N
N
Tamura
M
M
Okazaki
M
M
Delisle
B P
BP
January
C T
CT
Otsuji
Y
Y
D000935:Antifungal Agents / Q000008:administration & dosage / Q000009:adverse effects* / Q000494:pharmacology; D001665:Binding Sites; D015153:Blotting, Western; D002460:Cell Line; D004305:Dose-Response Relationship, Drug; D004594:Electrophysiology; D051638:Ether-A-Go-Go Potassium Channels / Q000037:antagonists & inhibitors*; D005434:Flow Cytometry; D006801:Humans; D007654:Ketoconazole / Q000008:administration & dosage / Q000009:adverse effects* / Q000494:pharmacology; D008133:Long QT Syndrome / Q000139:chemically induced*; D009124:Muscle Proteins / Q000378:metabolism; D009154:Mutation; D062554:NAV1.5 Voltage-Gated Sodium Channel; D018408:Patch-Clamp Techniques; D021381:Protein Transport / Q000187:drug effects; D015222:Sodium Channels / Q000378:metabolism; D013997:Time Factors
D000935:Antifungal Agents; D051638:Ether-A-Go-Go Potassium Channels; D009124:Muscle Proteins; D062554:NAV1.5 Voltage-Gated Sodium Channel; C568320:SCN5A protein, human; D015222:Sodium Channels; D007654:Ketoconazole