PERK modulation, with GSK2606414, Sephin1 or salubrinal, failed to produce therapeutic benefits in the SOD1G93A mouse model of ALS.

Amyotrophic lateral sclerosis (ALS) has been linked to overactivity of the protein kinase RNA-like ER kinase (PERK) branch of the unfolded protein response (UPR) pathway, both in ALS patients and mouse models. However, attempts to pharmacologically modulate PERK for therapeutic benefit have yielded inconsistent and often conflicting results. This study sought to address these discrepancies by comprehensively evaluating three commonly used, CNS-penetrant, PERK modulators (GSK2606414, salubrinal, and Sephin1) in the same experimental models, with the goal of assessing the viability of targeting the PERK pathway as a therapeutic strategy for ALS. To achieve this goal, a tunicamycin-challenge assay was developed using wild-type mice to monitor changes in liver UPR gene expression in response to PERK pathway modulation. Subsequently, multiple dosing regimens of each PERK modulator were tested in standardized, well-powered, gender-matched, and litter-matched survival efficacy studies using the SOD1G93A mouse model of ALS. The alpha-2-adrenergic receptor agonist clonidine was also tested to elucidate the results obtained from the Sephin1, and of the previously reported guanabenz studies, by comparing the effects of presence or absence of α-2 agonism. The results revealed that targeting PERK may not be an ideal approach for ALS treatment. Inhibiting PERK with GSK2606414 or activating it with salubrinal did not confer therapeutic benefits. While Sephin1 showed some promising therapeutic effects, it appears that these outcomes were mediated through PERK-independent mechanisms. Clonidine also produced some favorable therapeutic effects, which were unexpected and not linked to the UPR. In conclusion, this study highlights the challenges of pharmacologically targeting PERK for therapeutic purposes in the SOD1G93A mouse model and suggests that exploring other targets within, and outside, the UPR may be more promising avenues for ALS treatment.

SOD1-positive aggregate accumulation in the CNS predicts slower disease progression and increased longevity in a mutant SOD1 mouse model of ALS.

Non-natively folded variants of superoxide dismutase 1 (SOD1) are thought to contribute to the pathogenesis of familial amyotrophic lateral sclerosis (ALS), however the relative toxicities of these variants are controversial. Here, we aimed to decipher the relationships between the different SOD1 variants (aggregated, soluble misfolded, soluble total) and the clinical presentation of ALS in the SOD1G93A mouse. Using a multi-approach strategy, we found that the CNS regions least affected by disease had the most aggregated SOD1. We also found that the levels of aggregated SOD1 in the spinal cord were inversely correlated with the disease progression. Conversely, in the most affected regions, we observed that there was a high soluble misfolded/soluble total SOD1 ratio. Taken together, these findings suggest that soluble misfolded SOD1 may be the disease driver in ALS, whereas aggregated SOD1 may serve to sequester the toxic species acting in a neuroprotective fashion.

CuATSM efficacy is independently replicated in a SOD1 mouse model of ALS while unmetallated ATSM therapy fails to reveal benefits.

A copper chelator known as diacetylbis(N(4)-methylthiosemicarbazonato) copper II (CuATSM), has been reported to be efficacious in multiple transgenic SOD1 models of amyotrophic lateral sclerosis (ALS), a fatal neurodegenerative disorder affecting motor neurons. Here we report that we also observed CuATSM efficacy on disease onset and progression in a standardized litter-matched and gender-balanced efficacy study using B6SJL-SOD1G93A/1Gur mice. We also report improved survival trends with CuATSM treatment. In addition, we report a lack of efficacy by unmetallated ATSM in the same model using the same standardized study design. These results add to existing evidence supporting an efficacious role for copper delivery using chaperone molecules in mouse models of ALS.

A Quick Phenotypic Neurological Scoring System for Evaluating Disease Progression in the SOD1-G93A Mouse Model of ALS.

The SOD1-G93A transgenic mouse is the most widely used animal model of amyotrophic lateral sclerosis (ALS). At ALS TDI we developed a phenotypic screening protocol, demonstrated in video herein, which reliably assesses the neuromuscular function of SOD1-G93A mice in a quick manner. This protocol encompasses a simple neurological scoring system (NeuroScore) designed to assess hindlimb function. NeuroScore is focused on hindlimb function because hindlimb deficits are the earliest reported neurological sign of disease in SOD1-G93A mice. The protocol developed by ALS TDI provides an unbiased assessment of onset of paresis (slight or partial paralysis), progression and severity of paralysis and it is sensitive enough to identify drug-induced changes in disease progression. In this report, the combination of a detailed manuscript with video minimizes scoring ambiguities and inter-experimenter variability thus allowing for the protocol to be adopted by other laboratories and enabling comparisons between studies taking place at different settings. We believe that this video protocol can serve as an excellent training tool for present and future ALS researchers.

Guanabenz Treatment Accelerates Disease in a Mutant SOD1 Mouse Model of ALS.

Amyotrophic lateral sclerosis (ALS) is a progressive neurodegenerative disease characterized by loss of motor neurons. The mechanisms leading to motor neuron degeneration in ALS are unclear. However, there is evidence for involvement of endoplasmic reticulum (ER) stress and the unfolded protein response (UPR) in ALS, notably in mutant SOD1 mediated models of ALS. Stress induced phosphorylation of the eIF2 alpha subunit by eukaryotic translation initiation factor 2-alpha kinase 3 Perk activates the UPR. Guanabenz is a centrally acting alpha2 adrenergic receptor agonist shown to interact with a regulatory subunit of the protein phosphatase, Pp1/Gadd34, and selectively disrupt the dephosphorylation of the alpha subunit of eukaryotic initiation factor 2 (eif2alpha). Here we demonstrate that guanabenz is protective in fibroblasts expressing G93A mutant SOD1 when they are exposed to tunicamycin mediated ER stress. However, in contrast to other reports, guanabenz treatment accelerated ALS-like disease progression in a strain of mutant SOD1 transgenic ALS mice. This study highlights challenges of pharmacological interventions of cellular stress responses in whole animal models of ALS.

Dexpramipexole is ineffective in two models of ALS related neurodegeneration.

Treatment options for people living with amyotrophic lateral sclerosis (ALS) are limited and ineffective. Recently, dexpramipexole (RPPX) was advanced into human ALS clinical trials. In the current studies, we investigated RPPX in two parallel screening systems: 1) appropriately powered, sibling-matched, gender-balanced survival efficacy screening in high-copy B6-SJL-SOD1G93A/Gur1 mice, and 2) high-content neuronal survival screening in primary rat cortical neurons transfected with wild-type human TDP43 or mutant human TDP43. In both cases, we exposed the test systems to RPPX levels approximating those achieved in human Phase II clinical investigations. In SOD1G93A mice, no effect was observed on neuromotor disease progression or survival. In primary cortical neurons transfected with either mutant or wild-type human TDP43, a marginally significant improvement in a single indicator of neuronal survival was observed, and only at the 10 µM RPPX treatment. These systems reflect both mutant SOD1- and TDP43-mediated forms of neurodegeneration. The systems also reflect both complex non-cell autonomous and neuronal cell autonomous disease mechanisms. The results of these experiments, taken in context with results produced by other molecules tested in both screening systems, do not argue positively for further study of RPPX in ALS.

C57BL/6J congenic Prp-TDP43A315T mice develop progressive neurodegeneration in the myenteric plexus of the colon without exhibiting key features of ALS.

ALS therapy development has been hindered by the lack of rodent animal models. The discovery of TDP-43, a transcription factor that accumulates in the cytoplasm of motor neurons (MNs) in most cases of ALS, prompted attempts to develop TDP-43-based models of the disease. The current study sought to examine, in extensive detail, the emerging disease phenotype of a transgenic mouse model that overexpresses a mutant human TDP-43 (hTDP-43) gene under mouse prion promoter control. Careful attention was given to ALS-like characteristics to determine the appropriateness of this model for testing therapies for ALS. In light of previous reports that gastrointestinal (GI) dysfunction is responsible for early death in these mice, gut immunohistochemistry (IHC) and longitudinal gut motility assays were used to identify the onset and the progression of these defects. IHC studies revealed that site-specific overexpression of the hTDP-43 transgene in colonic myenteric plexes resulted in progressive neurodegeneration in this region. This change was associated with progressively reduced GI motility, culminating in frank stasis that was primarily responsible for decreasing longevity in these mice. The disease phenotype was gender- and genetic background-dependent, with congenic C57BL/6J male mice exhibiting the most aggressive form of the disease. Spinal cord IHC revealed ubiquitin-positive inclusions, but not TDP-43 aggregates, in the cytoplasm of MNs. Neither gender exhibited compelling ALS-like neuromuscular deficits, irrespective of age. While this model may be useful for studying GI tract neurodegeneration, in its present state it does not display a phenotype suitable for testing ALS therapeutics.

Treatment of shoulder pain utilizing mechanical diagnosis and therapy principles.

This case report describes the effectiveness of mechanical diagnosis and therapy (MDT) in the management of a patient referred with a diagnosis of shoulder tendonitis. The patient was a 56-year-old male with a 3-month history of left anterior shoulder pain. Upon initial assessment, he presented with a positive open-can test, lift-off test, and Hawkins-Kennedy impingement test. A MDT assessment quickly ruled out cervical involvement and identified a loss of end-range shoulder mobility and pain during active shoulder movement. After the patient underwent a repeated movement examination and treatment based on responses to end-range movements over three visits, his shoulder pain was abolished and motion was fully restored. Despite having positive rotator cuff and impingement signs, this patient was effectively treated with repeated end-range movements over a short period of 2 weeks. This case demonstrates that treatment based on MDT sub-classification principles may be an effective way to manage shoulder pain as it is in the spine.

From transcriptome analysis to therapeutic anti-CD40L treatment in the SOD1 model of amyotrophic lateral sclerosis.

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease characterized by progressive loss of motor neurons. Using unbiased transcript profiling in an ALS mouse model, we identified a role for the co-stimulatory pathway, a key regulator of immune responses. Furthermore, we observed that this pathway is upregulated in the blood of 56% of human patients with ALS. A therapy using a monoclonal antibody to CD40L was developed that slows weight loss, delays paralysis and extends survival in an ALS mouse model. This work demonstrates that unbiased transcript profiling can identify cellular pathways responsive to therapeutic intervention in a preclinical model of human disease.

No benefit from chronic lithium dosing in a sibling-matched, gender balanced, investigator-blinded trial using a standard mouse model of familial ALS.

BACKGROUND: In any animal model of human disease a positive control therapy that demonstrates efficacy in both the animal model and the human disease can validate the application of that animal model to the discovery of new therapeutics. Such a therapy has recently been reported by Fornai et al. using chronic lithium carbonate treatment and showing therapeutic efficacy in both the high-copy SOD1G93A mouse model of familial amyotrophic lateral sclerosis (ALS), and in human ALS patients. METHODOLOGY/PRINCIPAL FINDINGS: Seeking to verify this positive control therapy, we tested chronic lithium dosing in a sibling-matched, gender balanced, investigator-blinded trial using the high-copy (average 23 copies) SOD1G93A mouse (n = 27-28/group). Lithium-treated mice received single daily 36.9 mg/kg i.p. injections from 50 days of age through death. This dose delivered 1 mEq/kg (6.94 mg/kg/day lithium ions). Neurological disease severity score and body weight were determined daily during the dosing period. Age at onset of definitive disease and survival duration were recorded. Summary measures from individual body weight changes and neurological score progression, age at disease onset, and age at death were compared using Kaplan-Meier and Cox proportional hazards analysis. Our study did not show lithium efficacy by any measure. CONCLUSIONS/SIGNIFICANCE: Rigorous survival study design that includes sibling matching, gender balancing, investigator blinding, and transgene copy number verification for each experimental subject minimized the likelihood of attaining a false positive therapeutic effect in this standard animal model of familial ALS. Results from this study do not support taking lithium carbonate into human clinical trials for ALS.

Exercise self-efficacy, habitual physical activity, and fear of falling in patients with coronary heart disease.

The purpose of this study was to determine if a relationship exists between self-efficacy for physical activity and other pertinent factors in patients with coronary heart disease (CHD). A secondary purpose of this study was to determine if self-efficacy and exercise behavior are different in patients who report a fearing of falling (fallers) as compared to patients who do not report a fear of falling (non-fallers). This study included 50 patients who were admitted to the hospital for a CHD related diagnosis. Patients completed assessments of cardiac self-efficacy (Modified Barnason Efficacy Expectation Scale) and exercise behavior self-efficacy (Self Efficacy for Exercise Behavior Scale). In addition, the Physical Function subscale of the RAND 36-Item Health Survey and the Telephone Interview of Cognitive Function were used to characterize physical and cognitive function, respectively. Data analysis consisted of descriptive statistics, correlations, t-tests, and chi-square. Older patients reported higher levels of cardiac self-efficacy. Further, a positive correlation was found between cardiac self-efficacy and pre-hospitalization level of physical function. Patient income level and Self-efficacy for Exercise Behavior Resisting Relapse subscale scores were significantly correlated. A higher percent of fallers failed to meet minimum exercise guidelines as compared to non-fallers. It is important to identify the factors that are associated with exercise self-efficacy to improve health behavior adoption and adherence in patients with CHD.