巨大轴突性神经病（GAN）是一种由KLHL16基因突变引起的儿童神经退行性疾病。KLHL16基因编码着巨融素，起到了调节中间纤维（IF）周转的作用。过去的神经病理学研究以及当前研究中对尸体脑组织的检查揭示了星形细胞在GAN中的作用。为了开发一个临床相关的模型，我们将来自七个GAN患者的皮肤成纤维细胞重编程为多能干细胞（iPSCs），并用其生成了神经前体细胞（NPCs）、星形细胞和脑器官样结构。通过对携带G332R巨融素突变的一个患者细胞系进行CRISPR/Cas9基因编辑，获得了多个同基因对照克隆株。所有GAN iPSCs都缺乏巨融素并显示患者特异性的增加的波形蛋白表达。与同基因对照相比，GAN NPCs表达较低的巢蛋白，并且巢蛋白阳性细胞数量较少，但巢蛋白形态未受影响。GAN脑器官样结构中存在神经丝蛋白和GFAP的聚集。GAN iPSC-星形细胞显示出明显的高密度核周围波形蛋白和GFAP沉积以及异常的核形态。在超表达系统中，当存在波形蛋白时，GFAP寡聚体化和核周围聚集增强。具有大量核周围波形蛋白聚集的GAN患者细胞聚集了明显更多的核KLHL16 mRNA，与没有波形蛋白聚集的细胞相比。作为KLHL16突变的早期效应者，波形蛋白可能是GAN的一个潜在靶点。

Giant Axonal Neuropathy (GAN) is a pediatric neurodegenerative disease caused by KLHL16 mutations. KLHL16 encodes gigaxonin, which regulates intermediate filament (IF) turnover. Previous neuropathological studies and examination of postmortem brain tissue in the current study revealed involvement of astrocytes in GAN. To develop a clinically-relevant model, we reprogrammed skin fibroblasts from seven GAN patients to pluripotent stem cells (iPSCs), which were used to generate neural progenitor cells (NPCs), astrocytes, and brain organoids. Multiple isogenic control clones were derived via CRISPR/Cas9 gene editing of one patient line carrying the G332R gigaxonin mutation. All GAN iPSCs were deficient for gigaxonin and displayed patient-specific increased vimentin expression. GAN NPCs had lower nestin expression and fewer nestin-positive cells compared to isogenic controls, but nestin morphology was unaffected. GAN brain organoids were marked by the presence of neurofilament and GFAP aggregates. GAN iPSC-astrocytes displayed striking dense perinuclear vimentin and GFAP accumulations and abnormal nuclear morphology. In over-expression systems, GFAP oligomerization and perinuclear aggregation were augmented in the presence of vimentin. GAN patient cells with large perinuclear vimentin aggregates accumulated significantly more nuclear KLHL16 mRNA compared to cells without vimentin aggregates. As an early effector of KLHL16 mutations, vimentin may be a potential target in GAN.