Kumar Sourav, Nikelshparg Evelina, Pilátová Jana, Paul Ashim, Kumar Vijay, Koren Gil, Beck Roy, Jensen Henrik H, Segal Daniel
Shmunis School of Biomedicine and Cancer Research, George Wise Faculty of Life Sciences, Tel Aviv University, Tel Aviv 6997801, Israel.
Department of Life Sciences, Ben-Gurion University of the Negev, Beer Sheva 8855630, Israel.
ACS Nano. 2025 Jul 15;19(27):25180-25203. doi: 10.1021/acsnano.5c05498. Epub 2025 Jul 2.
Globoid cell leukodystrophy (GLD) is a rare hereditary inborn error of metabolism due to recessive mutations that cause loss of function of the enzyme galactosylceramidase (GALC). This results in the accumulation of the sphingolipids galactosylceramide (GalCer) and galactosylsphingosine (GalSph) in the lysosomes of neuronal cells. The accumulated GalCer and GalSph in cerebral macrophages of GLD patients are neurotoxic to oligodendrocytes and Schwann cells, leading to demyelination in the nervous system. The disease typically presents with infantile onset in the first six months of life and death by age 2. Here, we identified a supramolecular structure of GalCer and GalSph that may contribute to GLD pathology. Using biophysical assays commonly used for studying proteinaceous amyloids, e.g., amyloid-specific dyes, microscopical imaging, and a series of analytical methods (FTIR, PXRD, and SAXS), we demonstrate that both GalCer and GalSph can self-assemble in vitro into highly organized fibrils reminiscent of fibrils of amyloidogenic proteins. These fibrils exhibit significant cytotoxicity to both neuronal and oligodendroglial cells. Using an inhibitor of the GALC enzyme in cell culture to mimic the GLD pathophysiology, we could detect the accumulation of these fibrils in cells. We also observed that small molecules, which are bona fide inhibitors of proteinaceous amyloids, effectively mitigated the formation of the GalCer and GalSph fibrillar structures in vitro. Finally, the small molecule ameliorated the cytotoxic effects of the sphingolipid fibrils in SH-SY5Y cells, suggesting a potential avenue for therapeutic intervention in GLD orphan disease.
球形细胞脑白质营养不良（GLD）是一种罕见的遗传性先天性代谢缺陷病，由隐性突变导致半乳糖基神经酰胺酶（GALC）功能丧失引起。这导致鞘脂类半乳糖基神经酰胺（GalCer）和半乳糖基鞘氨醇（GalSph）在神经元细胞的溶酶体中积累。GLD患者脑巨噬细胞中积累的GalCer和GalSph对少突胶质细胞和施万细胞具有神经毒性，导致神经系统脱髓鞘。该疾病通常在出生后的头六个月内发病，两岁前死亡。在此，我们鉴定了一种GalCer和GalSph的超分子结构，其可能与GLD病理学有关。使用常用于研究蛋白质淀粉样蛋白的生物物理分析方法，例如淀粉样蛋白特异性染料、显微镜成像以及一系列分析方法（傅里叶变换红外光谱、粉末X射线衍射和小角X射线散射），我们证明GalCer和GalSph在体外均可自组装成高度有序的纤维，类似于淀粉样蛋白生成蛋白的纤维。这些纤维对神经元和少突胶质细胞均表现出显著的细胞毒性。在细胞培养中使用GALC酶抑制剂来模拟GLD病理生理学，我们能够检测到这些纤维在细胞中的积累。我们还观察到，作为蛋白质淀粉样蛋白的真正抑制剂的小分子，可有效减轻体外GalCer和GalSph纤维状结构的形成。最后，该小分子改善了鞘脂类纤维对SH-SY5Y细胞的细胞毒性作用，为GLD罕见病的治疗干预提供了一条潜在途径。
