抗癲癇藥物lacosamide與rufinamide對於電壓依賴型鈉離子通道之作用

Abstract

電壓控制型鈉離子通道對於誘發神經的興奮與動作電位的傳導，占有重要的地位。當腦部的神經興奮產生異常時，便會造成神經性疾病，例如癲癇的產生等等。而癲癇患者的治療方式，主要以給予抗電壓控制型鈉離子通道的藥物為主，像是carbamazepine, phenytoin 以及lamotrigine等典型的抗癲癇藥物，在過去的研究中，已被證實會與鈉離子通道的快速不活化態 (inactivation state)緩慢結合，並且相對於休息態的鈉離子通道，與快速不活化態有較高的親和力。另外，在過去的研究中，也發現到lacosamide (LCM)，此種較為新型的抗癲癇藥物，對於鈉離子通道作用有別於過去的典型藥物，並被提出可能會選擇性的結合到鈉離子通道的慢速不活化態。rufinamide (RFM)在臨床上主要用來治療LGS疾病與癲癇，而過去的研究主要著重在臨床以及動物實驗的研究上，較少提及此藥物對於鈉離子通道的分子作用機制。本篇論文利用多種實驗方法探討LCM與RFM兩種藥物主要是結合在鈉離子通道的快速不活化態抑或是作用於慢速不活化態。首先，LCM抑制鈉電流具有濃度依賴性與電壓依賴性，並且不同電壓下的抑制效果可由一對一結合曲線描繪，並且得到LCM對於不活化態之鈉離子通道的直觀解離常數(KI)為約150 μM。由LCM對鈉離子通道的結合速率常數與脫離速率常數所得的KI值可得到相似的結果，約140 μM。在不同濃度的LCM對電壓依賴不活化曲線中，得到KI值為約125 μM。因此可得LCM對於不活化態之鈉離子通道的親和力為125-150 μM，大於對休息態之鈉離子通道約50-60倍。另外，在比較給予長時間-80 mV或-10 mV的去極化後，LCM對鈉離子通道相較於控制組對前者有較慢回復至休息態之趨勢，對後者則無，表示LCM主要是對鈉離子通道的快速不活化態作用，而非慢速不活化態。同樣的，我們也對RFM以相同的實驗去驗證，得到RFM對於不活化態鈉離子通道的親和力為30-180 μM，大於對休息態的鈉離子通道約60-360倍。由此可得LCM與RFM對於鈉離子通道的快速不活化態具有相近的結合力，並且具有電壓依賴性。而在結合速率方面，RFM對於鈉離子通道之結合速率常數約為10300 M-1s-1，大於LCM對於鈉離子通道之結合速率約20倍。這些結果顯示，LCM與RFM皆是對於電壓依賴型鈉離子通道的不活化態作用，並且有相近的親和力，但RFM較LCM對於鈉離子通道之結合速率快非常多。因此，RFM相較LCM對於抑制鈉離子通道或許會有更快發生的效果。

Voltage-gated sodium channels play an important role in the genesis of action potential, and the cellular excitability. Epileptic seizure is a prototypical neurological disorder, characterized by abnormal neuronal excitability. Currently, quite a few anticonvulsants in clinical practice are targeted on voltage-gated sodium channels, such as carbamazepine, phenytoin and lamotrigine. These “classic’ anticonvulsants have been shown to bind to the fast-inactivated state rather than resting state of the Na+ channel. On the other hand, a newer anticonvulsant, lacosamide (LCM), was suggested to selectively bound to the slow-inactivated state of channel. Moreover, the other newer anticonvulsant, rufinamide (RFM), is used to treat Lennox-Gastaut syndrome in clinical practice. Because the molecular mechanisms of action of LCM and RFM have remained unclear, we used different experimental protocols to investigate the actions of LCM and RFM on the Na+ channel. LCM dose-dependently inhibited more Na+ currents at more depolarized holding potentials. The inhibitory effect could be described by one-to-one binding curves at various holding potentials, which yielded an apparent dissociation constant of ~150 μM for LCM binding to the inactivated channels (KI). A similar value of KI (~140 μM) was derived from the unbinding and binding rate constants. In addition, a KI of ~125 μM was also derived from the LCM concentration-dependent shift of the inactivation curve. Therefore, the affinity of LCM to inactivated Na+ channels was 125-150 μM, at least 50-60 times stronger than LCM binding to the resting channels. On the other hand, the recovery of LCM-bound inactivated Na+ channels was slower than normal (drug-free) after a prolonged depolarization for 18 sec at -80 mV but not at -10 mV, which indicates that LCM chiefly binds to the fast-inactivated state rather than the slow inactivated state. Similar investigations on RFM revealed that the dissociation constant of RFM binding to the fast inactivated Na+ channels is also around 30-180 μM, much stronger than binding to the resting state by 60-360 times. The macroscopic binding rate constant of RFM (10300 M-1s-1), however, is evidently faster than that of LCM (500 M-1s-1). In conclusion, these results indicate that LCM and RFM have significant effects on the fast-inactivated state of sodium channels, and similar affinity, but the binding rate of RFM is faster much more than LCM.