Synthesis and Evaluation of Ivacaftor Derivatives with Reduced Lipophilicity

Mutations in the
unique ATP-binding cassette anion channel, the
cystic fibrosis conductance regulator (CFTR), lead to the inherited
fatal disease known as cystic fibrosis (CF). Ivacaftor enhances channel
gating of CFTR by stabilizing its open state and has been approved
as monotherapy for CF patients with CFTR gating mutations (e.g., G551D)
and as part of combination therapy with lumacaftor for CFTR folding
mutations (e.g., ΔF508). However, in the latter context, ivacaftor
may destabilize folding-rescued ΔF508-CFTR and membrane-associated
proteins and attenuate lumacaftor pharmacotherapy. Here, we tested
the hypothesis that the high lipophilicity of ivacaftor may contribute
to this effect. We describe the synthesis of three glutamic acid ivacaftor
derivatives with reduced lipophilicity that bear different charges
at neutral pH (compounds 2, 3, 4). In a cellular ion flux assay, all three restored G551D-CFTR channel
activity at comparable or better levels than ivacaftor. Furthermore,
unlike ivacaftor, compound 3 did not attenuate levels
of folding-rescued ΔF508 at the cell surface. Molecular modeling
predicts that the increased polarity of compound 3 allows
engagement with polar amino acids present in the binding pocket with
hydrogen bonding and ionic interactions, which are collectively higher
in strength as compared to hydrophobic interactions that stabilize
ivacaftor. Overall, the data suggests that reduced lipophilicity may
improve the efficacy of this class of CFTR potentiators when used
for folding-rescued ΔF508-CFTR.

囊性纤维化跨膜传导调节蛋白（cystic fibrosis conductance regulator, CFTR）属于一类独特的ATP结合盒阴离子通道，其编码基因发生突变会引发遗传性致死疾病囊性纤维化（cystic fibrosis, CF）。依伐卡托（Ivacaftor）可通过稳定CFTR的开放状态增强其通道门控功能，目前已被批准作为单一疗法，用于携带CFTR门控突变（如G551D）的囊性纤维化患者；同时也可与卢马卡托（lumacaftor）联合使用，用于携带CFTR折叠突变（如ΔF508）的患者。然而在联合治疗的场景中，依伐卡托可能会破坏经折叠拯救的ΔF508-CFTR以及膜相关蛋白的稳定性，并削弱卢马卡托的药效。本研究针对“依伐卡托的高脂溶性可能是导致该现象的原因”这一假说展开验证。我们合成了三种脂溶性降低的谷氨酸-依伐卡托衍生物，这三种化合物在中性pH环境下带有不同电荷，分别记为化合物2、3、4。在细胞离子通量检测实验中，三种衍生物均能恢复G551D-CFTR的通道活性，其效果与依伐卡托相当甚至更优。此外，与依伐卡托不同的是，化合物3不会降低经折叠拯救的ΔF508-CFTR在细胞表面的表达水平。分子建模结果显示，化合物3的极性更强，可通过氢键与离子相互作用与结合口袋内的极性氨基酸结合，这类相互作用的整体强度高于稳定依伐卡托的疏水相互作用。综上，本研究数据表明，若将脂溶性降低的CFTR增效剂类化合物用于经折叠拯救的ΔF508-CFTR治疗，或可提升其临床疗效。