# Axonal pathogenesis of human iPSC-derived motor neurons

> **NIH NIH R21** · JOHNS HOPKINS UNIVERSITY · 2022 · $450,313

## Abstract

Project Summary:
Reprogramming adult cells has made it possible to differentiate patient-specific neurons from induced
pluripotent stem cells (iPSCs). These patient-derived neurons have become invaluable in the investigation of
molecular mechanisms of neurodegeneration and identification of potential therapeutic targets. Amyotrophic
Lateral Sclerosis (ALS) patient-derived spinal motor neurons have revealed insights into mutation-specific
pathogenesis, but previous studies have almost exclusively addressed deficits within motor neurons such as
stress granule formation, hyperexcitability, and reduced autophagy. However, the initial pathology of ALS, and
many other motor neuron diseases, begins at the distal axon and neuromuscular junction. Normal adult axons
contain thousands of diverse sets of mRNAs whose protein products are locally translated to maintain axonal
homeostasis and health. However, the expression of axonal mRNA in human motor neurons derived from
iPSCs harboring ALS-linked mutations are poorly understood. Additionally, iPSC-derived neuromuscular
synapses are rarely utilized as a system to interrogate the pathogenesis of axon degeneration, in part,
because of a lack of robust and systematic evaluations of neuromuscular synapses formed by different iPSC-
derived cells. The main questions that this application addresses are:
1) Are there differences in the expression of axonal mRNA between patient and healthy control iPSC-
derived human spinal motor neurons?
2) What are the effects of distinct disease-causing mutations on the formation and maintenance of
human muscle fiber innervations?
We hypothesize that ALS-causing mutations may reduce the abundance of locally translated axonal mRNA
whose protein products are involved in axonal health and function. We will subject axonal RNA extracted from
control and mutant samples to RNAseq, and we are collaborating with bioinformatic group at our institution to
probe for effected axonal pathways. We plan to take advantage of a microfluidic device platform that separate
neuronal cell bodies from axons/muscle fibers, permitting us to obtain pure axonal mRNA and create mature
human-derived neuromuscular synapses. We will test human cell lines containing distinct mutations: SOD1
and C9orf72 repeat expansions, and we will first examine SOD1A4V lines and C9orf72 lines for which isogenic
controls have been created. In addition, we have access to cell lines in which the genetic cause of disease has
not been determined (sporadic ALS) through the Johns Hopkins ALS center and Answer ALS that we can use
for future studies.

## Key facts

- **NIH application ID:** 10604850
- **Project number:** 1R21NS130900-01
- **Recipient organization:** JOHNS HOPKINS UNIVERSITY
- **Principal Investigator:** Mohamed H Farah
- **Activity code:** R21 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2022
- **Award amount:** $450,313
- **Award type:** 1
- **Project period:** 2022-09-19 → 2024-08-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10604850

## Citation

> US National Institutes of Health, RePORTER application 10604850, Axonal pathogenesis of human iPSC-derived motor neurons (1R21NS130900-01). Retrieved via AI Analytics 2026-05-26 from https://api.ai-analytics.org/grant/nih/10604850. Licensed CC0.

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