Human Amyotrophic Lateral Sclerosis iPSC Model
Amyotrophic lateral sclerosis, ALS, also called Lou Gehrig's Disease, is a fatal, late-onset disease with severe diagnosis and no cure today. ALS targets motor neurons, causing their loss and, eventually, to the damage of motor function and muscle movement. ALS occurs as a bulbar or spinal ALS.ALS is a rare disease with no clear etiology. 10% of patients have a familial hereditary form of ALS - familial ALS, fALS -but most patients, 90%, suffer from sporadic ALS. Sporadic and familial forms are clinically identical.
Familial Forms, SOD1, C9ORF72, and TDP43, of Amyotrophic Lateral Sclerosis
Mutations of C9orf72, SOD1, TDP43, and others cause familial ALS, which are essential and valuable for ALS disease modeling for phenotypic screening.
The mutations C9orf72, SOD1, and TDP43 cover different pathological mechanisms of ALS so that phenotypic assays with these other mutated iPSC-derived motor neurons will gain their predictive power.
The C9orf72 gene is implicated in ALS through the repetition of GGGGCC expansion in non-coding regions with effects on RNA binding and autophagy regulation. The C9orf72 mutation is also causing, in some cases, frontal temporal dementia.
The mutations of SOD1 affect the enzyme Cu/Zn-superoxide dismutase 1. These mutations cause the degeneration of motor neurons by oxidative stress, protein aggregates, mitochondrial defects, and glutamate excitotoxicity.
The TAR DNA-binding protein 43 (TDP-43) is another mutation that causes fALS.
We have detected in C9orf72 cells mislocalized proteins of TDP-43:
Spinal motor neurons with TDP-43 in somas.
C9orf72 spinal motor neuron cells with TDP-43 in somas and axons.
Neurodegenerative mechanisms in ALS are very diverse, which explains the difficulties of developing new treatments for this disease. NeuroProof offers different ALS iPSC-derived disease models for testing treatments in other disease mechanisms. Astrocytes and microglia are also implicated in ALS disease mechanisms. For this, we can perform co-culture or tri-culture assays.
Human iPSC-Derived Spinal Motor Neuron Cells with SOD1, and C9orf72 Mutation, and Healthy Control Neuronal Activity
We present the neuronal activity for the three human spinal motor neuron cell lines cultivated on microelectrode arrays. These motor neurons form spontaneous activity patterns, see neural stem cells.
The cell lines with the SOD1, C9orf72, and TDP-43 mutation reveal disturbed excitatory-inhibitory balance, E/I balance. In particular, iPSC-derived motoneuron cultures with a SOD1, C9orf72, and TDP-43 mutation show a hyper-excitation compared to the healthy control described above.
Image: Spike train parameter comparison of a SOD1-D90A cell line, its isogenic control, and a C9orf72 cell line, demonstrating a highly hyper-excited activity pattern of the ALS mutated cell lines.
Description of an example with a SOD1-D90A mutation ALS Rescue Assay
As an example, we illustrate a rescue assay that rescues the electrical activity pattern of a mutated SOD1 iPSC-derived motor neuron cell line back to a healthy phenotype, the activity of its isogenic control cell line. We recorded the spike train data on day 11 in vitro.
Spike train parameters were calculated with NeuroProof's proprietary spike train analysis tools. With a specific projection of the spike train parameters into a one-dimensional effect score, it is then possible to assess the putative therapeutic effect of a compound.
Image: Example of a compound effect in an ALS rescue assay. The reversion of a compound is assessed from the diseased cell line activity normalized at one to the healthy cell line normalized at zero.
Further readouts, such as detecting TDP-43 misfolding or as a survival assay, can complement the functional phenotypic readouts of the MEA recordings. NeuroProof offers assays with different ALS cell lines, which can be combined with astrocytes or microglia.
NeuroProof offers tests with the following ALS cell lines:
|C9orf72||patient cell line|
The cells can also be cultured for more extended periods. In these cells cultured for a longer period, NF-L, neurofilament light chain, can be detected as a clinical marker for ALS.
Customers can order a microelectrode array standalone readout or a combination with other readouts, such as:
- Imaging readouts of immunostained cells as for TDP-43, for example.
- Molecular readouts for cell viability or neurofilament senescence.
NeuroProof is continuously expanding its assay portfolio, asking us for potential new ALS cell lines with familial or sporadic forms and co-cultures with astrocytes or/and microglia. NeuroProof performs integrated projects, from sourcing diseased cell lines, cell differentiation, and assay development to compound screening campaigns.