Shen Liwei, Ye Xiaofei, Wang Xiaocui, Song Conglei, Yang Bin
Department of Neurology, Anhui Provincial Children's Hospital/Children's Hospital of Fudan University (Affiliated Anhui Branch), Hefei, China.
Mol Genet Genomic Med. 2025 Apr;13(4):e70099. doi: 10.1002/mgg3.70099.
Biallelic variants of COL18A1 cause Knobloch syndrome (KNO), a rare genetic disorder, characterized by oculopathy and structural defects. Recently, several studies have suggested that novel de novo missense variants in PAK2 may be associated with KNO; however, there are few case reports. This study aimed to investigate a patient with KNO who initially presented with seizures and expand the PAK2 genotype and phenotype spectrum.
This study included a Chinese family with a proband who primarily presented with epilepsy and developmental delay. Whole-exome sequencing and Sanger sequencing were performed to analyze potential variants. Structural modeling was performed to assess the impact of the variant on the protein structure. In vitro, a mutant plasmid was constructed and transfected into 293T cells to conduct phosphorylation assays, and phosphorylation levels at Ser141 of PAK2 were assessed. The PAK kinase inhibitor FRAX597 was used to confirm the specificity of the western blot results.
A de novo variant of PAK2 gene, NM_002577.4: c.1049G>A (p.Arg350Lys) was found in the patient but not in his parents or sister. This variant was found to be located in the kinase domain and may alter the hydrogen-bond network, potentially affecting kinase activity. In vitro functional experiments demonstrated that the variant may lead to reduced protein levels. Moreover, Western blot analysis showed a significant decrease in the phosphorylation level at Ser141 compared to the wild-type plasmid, indicating that the variant may lead to decreased PAK2 phosphorylation levels.
The clinical manifestations in this patient may be associated with a novel PAK2 variant, and the atypical presentation of KNO suggests that PAK2-related KNO may have a broader phenotypic spectrum.
COL18A1双等位基因变异导致诺布洛克综合征（KNO），这是一种罕见的遗传性疾病，其特征为眼病和结构缺陷。最近，多项研究表明，PAK2基因中的新型新生错义变异可能与KNO相关；然而，病例报告较少。本研究旨在调查一名最初表现为癫痫发作的KNO患者，并扩展PAK2基因的基因型和表型谱。
本研究纳入了一个中国家庭，先证者主要表现为癫痫和发育迟缓。进行全外显子组测序和桑格测序以分析潜在变异。进行结构建模以评估该变异对蛋白质结构的影响。在体外，构建突变体质粒并转染至293T细胞中进行磷酸化测定，并评估PAK2第141位丝氨酸的磷酸化水平。使用PAK激酶抑制剂FRAX597来确认蛋白质印迹结果的特异性。
在患者中发现了PAK2基因的一个新生变异，NM_002577.4:c.1049G>A（p.Arg350Lys），但其父母和姐姐未发现该变异。该变异位于激酶结构域，可能会改变氢键网络，潜在影响激酶活性。体外功能实验表明，该变异可能导致蛋白质水平降低。此外，蛋白质印迹分析显示，与野生型质粒相比，第141位丝氨酸的磷酸化水平显著降低，表明该变异可能导致PAK2磷酸化水平降低。
该患者的临床表现可能与一种新型PAK2变异有关，KNO的非典型表现提示PAK2相关的KNO可能具有更广泛的表型谱。
