# A Dual Strategy—In Vitro and In Silico—To Evaluate Human Antitetanus mAbs Addressing Their Potential Protective Action on TeNT Endocytosis in Primary Rat Neuronal Cells

## TL;DR

Researchers tested human antibodies against tetanus toxin to see if they can block its entry into nerve cells, using both lab experiments and computer modeling.

## Contribution

The study introduces a dual in vitro and in silico approach to evaluate human monoclonal antibodies against tetanus neurotoxin endocytosis.

## Key findings

- Two human monoclonal antibodies inhibited tetanus neurotoxin (TeNT) internalization in rat neuronal cells by over 50%.
- Molecular docking confirmed antibody binding to the HCR/T domain of TeNT and identified key residues for epitope formation.
- The Synaptic Vesicle Glycoprotein II (SV2) was confirmed as a mediator in TeNT endocytosis.

## Abstract

Tetanus disease, caused by C. tetani, starts with wounds or mucous layer contact. Prevented by vaccination, the lack of booster shots throughout life requires prophylactic treatment in case of accidents. The incidence of tetanus is high in underdeveloped countries, requiring the administration of antitetanus antibodies, usually derived from immunized horses or humans. Heterologous sera represent risks such as serum sickness. Human sera can carry unknown viruses. In the search for human monoclonal antibodies (mAbs) against TeNT (Tetanus Neurotoxin), we previously identified a panel of mAbs derived from B-cell sorting, selecting two nonrelated ones that binded to the C-terminal domain of TeNT (HCR/T), inhibiting its interaction with the cellular receptor ganglioside GT1b. Here, we present the results of cellular assays and molecular docking tools. TeNT internalization in neurons is prevented by more than 50% in neonatal rat spinal cord cells, determined by quantitative analysis of immunofluorescence punctate staining of Alexa Fluor 647 conjugated to TeNT. We also confirmed the mediator role of the Synaptic Vesicle Glycoprotein II (SV2) in TeNT endocytosis. The molecular docking assays to predict potential TeNT epitopes showed the binding of both antibodies to the HCR/T domain. A higher incidence was found between N1153 and W1297 when evaluating candidate residues for conformational epitope.