CuATSM effectively ameliorates ALS patient astrocyte‐mediated motor neuron toxicity in human in vitro models of amyotrophic lateral sclerosis

Abstract

Patient diversity and unknown disease cause are major challenges for drug development and clinical trial design for amyotrophic lateral sclerosis (ALS). Transgenic animal models do not adequately reflect the heterogeneity of ALS. Direct reprogramming of patient fibroblasts to neuronal progenitor cells and subsequent differentiation into patient astrocytes allows rapid generation of disease relevant cell types. Thus, this methodology can facilitate compound testing in a diverse genetic background resulting in a more representative population for therapeutic evaluation. Here, we used established co‐culture assays with motor neurons and reprogrammed patient skin‐derived astrocytes (iAs) to evaluate the effects of (SP‐4‐2)‐[[2,2’‐(1,2‐dimethyl‐1,2‐ethanediylidene)bis[N‐methylhydrazinecarbothioamidato‐κN2,κS]](2‐)]‐copper (CuATSM), currently in clinical trial for ALS in Australia. Pretreatment of iAs with CuATSM had a differential effect on neuronal survival following co‐culture with healthy motor neurons. Using this assay, we identified responding and non‐responding cell lines for both sporadic and familial ALS (mutant SOD1 and C9ORF72). Importantly, elevated mitochondrial respiration was the common denominator in all CuATSM‐responders, a metabolic phenotype not observed in non‐responders. Pre‐treatment of iAs with CuATSM restored mitochondrial activity to levels comparable to healthy controls. Hence, this metabolic parameter might allow selection of patient subpopulations best suited for CuATSM treatment. Moreover, CuATSM might have additional therapeutic value for mitochondrial disorders. Enhanced understanding of patient‐specific cellular and molecular profiles could help improve clinical trial design in the future.

Treatment with the copper compound CuATSM has no significant effect on motor neuronal pathology in patients with ALS

“…Our results revealed no significant difference in neuron density or TDP burden in patients that had received CuATSM compared with patients that had not. In contrast to that seen in preclinical models [3–6, 8, 22, 23], SOD1 immunoreactivity and GFAP expression were similar in CuATSM and non‐CuATSM treatment groups. However, patients treated with CuATSM demonstrated p62‐immunoreactive astrocytes in the motor cortex and reduced Iba1 density in the spinal cord.…”

“…P62 is an autophagy substrate that plays a critical role in aggregate degeneration [ 24 ] and was observed here in motor cortical astrocytes of CuATSM‐treated patients only, where it filled the astrocyte cytoplasm in a similar way to GFAP (Figure 1 ). Although p62‐immunopositive astrocytes have not been described in postmortem tissue of patients with ALS, elevated p62 levels have been identified in reprogrammed skin‐derived astrocytes from patients with sporadic, SOD1 and C9ORF72 ALS [ 23 , 25 ]. Importantly, no relationship between astroglial sequestosome activity with CuATSM treatment was found in patient cell lines [ 23 ], suggesting that the p62‐astrocytes observed here do not reflect a therapeutic response to CuATSM.…”

“…Although p62‐immunopositive astrocytes have not been described in postmortem tissue of patients with ALS, elevated p62 levels have been identified in reprogrammed skin‐derived astrocytes from patients with sporadic, SOD1 and C9ORF72 ALS [ 23 , 25 ]. Importantly, no relationship between astroglial sequestosome activity with CuATSM treatment was found in patient cell lines [ 23 ], suggesting that the p62‐astrocytes observed here do not reflect a therapeutic response to CuATSM. Further to this, in contrast to the reduced astrocytic activity seen in SOD1 rodent models [ 3 , 4 , 5 , 6 , 7 , 8 ], no significant change in measures of GFAP was found with CuATSM here.…”

Novel human neurodevelopmental and neurodegenerative disease associated with IRF2BPL gene variants—mechanisms and therapeutic avenues

“…The literature suggests that CuATSM’s potential mechanisms involve the restoration of mitochondrial function, including the correction of increased fractionation and mislocalization of mitochondria observed in NEDAMSS patient cells ( Sinha Ray et al, 2022 ). The drug is safe for use in humans and is currently in clinical trials for hypoxic imaging, ALS treatment ( Dennys et al, 2023 ), and Parkinson’s disease ( Nikseresht et al, 2020 ; Sinha Ray et al, 2022 ). In the phase I clinical trial of ALS, the drug slowed down disease progression and improved the respiratory and cognitive function of the patients ( Sinha Ray et al, 2022 ).…”

Mechanisms of IRF2BPL-related disorders and identification of a potential therapeutic strategy

“…In our previous study involving coculture assays with ALS astrocytes, we had observed a similar loss of neuronal viability, and treatment with CuATSM drug significantly improved the survival in certain responding patient lines. 25 CuATSM is a small artificial drug currently being used in clinical trials for ALS 26,27 and Parkinson's disease (PD) (NCT03204929). [26][27][28] Interestingly, all patientderived iAs treated with 1 mM CuATSM for 4 consecutive days prior to neuron-glia coculture displayed significantly increased neuronal survival (Figures 3A and 3C).…”