Anti-SOD1 Nanobodies That Stabilize Misfolded SOD1 Proteins Also Promote Neurite Outgrowth in Mutant SOD1 Human Neurons

ALS-linked mutations induce aberrant conformations within the SOD1 protein that are thought to underlie the pathogenic mechanism of SOD1-mediated ALS. Although clinical trials are underway for gene silencing of SOD1, these approaches reduce both wild-type and mutated forms of SOD1. Here, we sought to develop anti-SOD1 nanobodies with selectivity for mutant and misfolded forms of human SOD1 over wild-type SOD1. Characterization of two anti-SOD1 nanobodies revealed that these biologics stabilize mutant SOD1 in vitro. Further, SOD1 expression levels were enhanced and the physiological subcellular localization of mutant SOD1 was restored upon co-expression of anti-SOD1 nanobodies in immortalized cells. In human motor neurons harboring the SOD1 A4V mutation, anti-SOD1 nanobody expression promoted neurite outgrowth, demonstrating a protective effect of anti-SOD1 nanobodies in otherwise unhealthy cells. In vitro assays revealed that an anti-SOD1 nanobody exhibited selectivity for human mutant SOD1 over endogenous murine SOD1, thus supporting the preclinical utility of anti-SOD1 nanobodies for testing in animal models of ALS. In sum, the anti-SOD1 nanobodies developed and presented herein represent viable biologics for further preclinical testing in human and mouse models of ALS.

Automated summary

ALS-linked mutations can cause aberrant conformations in the SOD1 protein, and this study aimed to develop anti-SOD1 nanobodies that specifically target mutant and misfolded forms of SOD1. The researchers found that these nanobodies can stabilize mutant SOD1, enhance its expression, and restore its subcellular localization. In addition, the nanobodies promote neurite outgrowth in human motor neurons harboring the SOD1 A4V mutation and exhibit selectivity for human mutant SOD1 over mouse SOD1 in vitro, supporting their potential for preclinical testing in animal models.