The molecular tweezer CLR01 inhibits aberrant superoxide dismutase 1 (SOD1) self-assembly in vitro and in the G93A-SOD1 mouse model of ALS

Malik, R. K.; Meng, Helen; Wongkongkathep, Piriya; Corrales, Christian I.; Sepanj, Niki; Atlasi, Ryan S.; Klärner, Frank‐Gerrit; Schräder, Thomas; Spencer, Melissa J.; Loo, Joseph A.; Wiedau, Martina; Bitan, Gal

doi:10.1074/jbc.ra118.005940

Cited by 36 publications

(51 citation statements)

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“…De‐metallated SOD1 was used for the aggregation assay as in this form SOD1 can easily be induced to aggregate by reducing its disulfide bonds (22‐24). SOD1 was incubated at a concentration of 40 µM in the absence or presence of 0.3, 1, 3, or 10 molar equivalents of each compound in 10 mM potassium phosphate, pH 7.4, containing 40 µM ThT and 50 mM Tris(2‐carboxyethyl)phosphine (TCEP) in a final volume of 100 µL (19). The aggregation assay was carried out in 96‐well, opaque‐walls, clear flat‐bottom wells, each containing a Teflon ball to assist with agitation.…”

Section: Methodsmentioning

confidence: 99%

Examination of SOD1 aggregation modulators and their effect on SOD1 enzymatic activity as a proxy for potential toxicity

et al. 2020

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Small‐molecule inhibitors of abnormal protein self‐assembly are promising candidates for developing therapy against proteinopathies. Such compounds have been examined primarily as inhibitors of amyloid β‐protein (Aβ), whereas testing of inhibitors of other amyloidogenic proteins has lagged behind. An important issue with screening compound libraries is that although an inhibitor suitable for therapy must be both effective and nontoxic, typical screening focuses on efficacy, whereas safety typically is tested at a later stage using cells and/or animals. In addition, typical thioflavin T (ThT)‐fluorescence‐based screens use the final fluorescence value as a readout, potentially missing important kinetic information. Here, we examined potential inhibitors of superoxide dismutase 1 (SOD1) using ThT‐fluorescence including the different phases of fluorescence change and added a parallel screen of SOD1 activity as a potential proxy for compound toxicity. Some compounds previously reported to inhibit other amyloidogenic proteins also inhibited SOD1 aggregation at low micromolar concentrations, whereas others were ineffective. Analysis of the lag phase and exponential slope added important information that could help exclude false‐positive or false‐negative results. SOD1 was highly resistant to inhibition of its activity, and therefore, did not have the necessary sensitivity to serve as a proxy for examining potential toxicity.

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Section: Methodsmentioning

confidence: 99%

Examination of SOD1 aggregation modulators and their effect on SOD1 enzymatic activity as a proxy for potential toxicity

et al. 2020

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“…For example, the mSOD1-G93A transgenic mouse model of ALS [which overexpresses the human SOD1 with the Gly-93-Ala (G93A) substitution], has been revealed to display reduced activity of mitochondrial complex I (Jung et al, 2002;Coussee et al, 2011). Interestingly, the discovery of the G93A mutation in the antioxidant enzyme SOD1 was the first known genetic cause of human ALS, and ∼160 different mutations affecting the binding of Cu and Zn to the redox center of SOD1 have been identified (Lovejoy and Guillemin, 2014;Malik et al, 2019). There is also much evidence that transition metals, especially Cu, Zn, and Fe, can mediate mitochondrial dysfunction, DNA damage, telomere shortening, and neurodegeneration (Almeida et al, 2006;Lovejoy and Guillemin, 2014;Bertoncini et al, 2016).…”

Section: Mitochondrial Dysfunction In Alsmentioning

confidence: 99%

Mitochondrial Dysfunction, Neurogenesis, and Epigenetics: Putative Implications for Amyotrophic Lateral Sclerosis Neurodegeneration and Treatment

et al. 2020

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Amyotrophic lateral sclerosis (ALS) is a progressive and devastating multifactorial neurodegenerative disorder. Although the pathogenesis of ALS is still not completely understood, numerous studies suggest that mitochondrial deregulation may be implicated in its onset and progression. Interestingly, mitochondrial deregulation has also been associated with changes in neural stem cells (NSC) proliferation, differentiation, and migration. In this review, we highlight the importance of mitochondrial function for neurogenesis, and how both processes are correlated and may contribute to the pathogenesis of ALS; we have focused primarily on preclinical data from animal models of ALS, since to date no studies have evaluated this link using human samples. As there is currently no cure and no effective therapy to counteract ALS, we have also discussed how improving neurogenic function by epigenetic modulation could benefit ALS. In support of this hypothesis, changes in histone deacetylation can alter mitochondrial function, which in turn might ameliorate cellular proliferation as well as neuronal differentiation and migration. We propose that modulation of epigenetics, mitochondrial function, and neurogenesis might provide new hope for ALS patients, and studies exploring these new territories are warranted in the near future.

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“…Experimental design and safety assessment of CLR01 treatment Previously, CLR01 was used to treat different animal models of proteinopathy using intracerebroventribular (32,36) or subcutaneous administration. In the latter route, the compound was administered either via osmotic minipumps (20,25,36,37), as was done here, or by daily injection, 2-7 days a week (38)(39)(40)(41). Here, we chose to use the less labor-intensive osmotic-minipump subcutaneous administration to answer the speci c question whether CLR01 treatment affected tau directly.…”

Section: Mouse Modelmentioning

confidence: 99%

The molecular tweezer CLR01 improves behavioral deficits and reduces tau pathology in P301S-tau transgenic mice

Siddique

et al. 2020

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Background: Molecular tweezers (MTs) are broad-spectrum inhibitors of abnormal protein aggregation. A lead MT, called CLR01, has been demonstrated to inhibit the aggregation and toxicity of multiple amyloidogenic proteins in vitro and in vivo. Previously, we evaluated the effect of CLR01 in the 3×Tg mouse model of Alzheimer’s disease, which overexpresses mutant human presenilin 1, amyloid β-protein precursor, and tau and found that subcutaneous administration of the compound for one month led to a robust reduction of amyloid plaques, neurofibrillary tangles, and microgliosis. CLR01 also has been demonstrated to inhibit tau aggregation in vitro and tau seeding in cell culture, yet because in Alzheimer’s disease (AD) and in the 3×Tg model, tau hyperphosphorylation and aggregation are thought to be downstream of Aβ insults, the study in this model left the question whether CLR01 affected tau in vivo directly or indirectly open.Methods: To determine if CLR01 could ameliorate tau pathology directly in vivo, we tested the compound similarly using the P301S-tau (line PS19) mouse model. Mice were administered 0.3- or 1.0-mg/Kg per day CLR01 and tested for muscle strength and behavioral deficits, including anxiety- and disinhibition-like behavior. Their brains then were analyzed by immunohistochemical and biochemical assays for pathological forms of tau, neurodegeneration, and glial pathology.Results: CLR01 treatment ameliorated muscle-strength deterioration, anxiety-, and disinhibition-like behavior. Improved phenotype was associated with decreased levels of pathologic tau forms, suggesting that CLR01 exerts a direct effect on tau in vivo. Limitations of the study included a relatively short treatment period of the mice at an age in which full pathology is not yet developed. In addition, high variability in this model lowered the statistical significance of the findings of some outcome measures.Conclusions: The findings suggest that CLR01 is a particularly attractive candidate for the treatment of AD because it targets simultaneously the two major pathogenic proteins instigating and propagating the disease, amyloid β-protein (Aβ) and tau, respectively. In addition, our study suggests that CLR01 can be used for the treatment of other tauopathies in the absence of amyloid pathology.

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The molecular tweezer CLR01 inhibits aberrant superoxide dismutase 1 (SOD1) self-assembly in vitro and in the G93A-SOD1 mouse model of ALS

Cited by 36 publications

References 51 publications

Examination of SOD1 aggregation modulators and their effect on SOD1 enzymatic activity as a proxy for potential toxicity

Examination of SOD1 aggregation modulators and their effect on SOD1 enzymatic activity as a proxy for potential toxicity

Mitochondrial Dysfunction, Neurogenesis, and Epigenetics: Putative Implications for Amyotrophic Lateral Sclerosis Neurodegeneration and Treatment

The molecular tweezer CLR01 improves behavioral deficits and reduces tau pathology in P301S-tau transgenic mice

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