A novel cell immunoassay to measure survival of motor neurons protein in blood cells

BACKGROUND: The motor neuron degenerative disease spinal muscular atrophy (SMA) is the leading genetic cause of infant mortality and is caused by mutations in the survival of motor neurons (SMN) gene that reduce the expression levels of the SMN protein. A major goal of current therapeutic approaches is to increase SMN levels in SMA patients. The purpose of this study was to develop a reliable assay to measure SMN protein levels from peripheral blood samples. METHODS: We developed a novel cell immunoassay to quantitatively measure SMN levels from peripheral blood mononuclear cells (PBMCs) using a single anti-SMN antibody. RESULTS: SMN levels determined by the cell immunoassay are comparable to levels determined by Western blot, but in contrast, the immunoassay does not involve cell lysis, requires a small amount of patient material, and can be done on a large number of samples simultaneously. SMN levels from PBMCs are not influenced by cell type heterogeneity. CONCLUSION: SMN levels measured from total PBMCs provide an important snapshot of SMN protein expression, which should be a useful aid in SMA diagnosis, and a surrogate marker of efficacy of treatment in SMA clinical trials

A call for transparent reporting to optimize the predictive value of preclinical research

The US National Institute of Neurological Disorders and Stroke convened major stakeholders in June 2012 to discuss how to improve the methodological reporting of animal studies in grant applications and publications. The main workshop recommendation is that at a minimum studies should report on sample-size estimation, whether and how animals were randomized, whether investigators were blind to the treatment, and the handling of data. We recognize that achieving a meaningful improvement in the quality of reporting will require a concerted effort by investigators, reviewers, funding agencies and journal editors. Requiring better reporting of animal studies will raise awareness of the importance of rigorous study design to accelerate scientific progress

The Survival of Motor Neurons Protein Determines the Capacity for snRNP Assembly: Biochemical Deficiency in Spinal Muscular Atrophy

Reduction of the survival of motor neurons (SMN) protein levels causes the motor neuron degenerative disease spinal muscular atrophy, the severity of which correlates with the extent of reduction in SMN. SMN, together with Gemins 2 to 7, forms a complex that functions in the assembly of small nuclear ribonucleoprotein particles (snRNPs). Complete depletion of the SMN complex from cell extracts abolishes snRNP assembly, the formation of heptameric Sm cores on snRNAs. However, what effect, if any, reduction of SMN protein levels, as occurs in spinal muscular atrophy patients, has on the capacity of cells to produce snRNPs is not known. To address this, we developed a sensitive and quantitative assay for snRNP assembly, the formation of high-salt- and heparin-resistant stable Sm cores, that is strictly dependent on the SMN complex. We show that the extent of Sm core assembly is directly proportional to the amount of SMN protein in cell extracts. Consistent with this, pulse-labeling experiments demonstrate a significant reduction in the rate of snRNP biogenesis in low-SMN cells. Furthermore, extracts of cells from spinal muscular atrophy patients have a lower capacity for snRNP assembly that corresponds directly to the reduced amount of SMN. Thus, SMN determines the capacity for snRNP biogenesis, and our findings provide evidence for a measurable deficiency in a biochemical activity in cells from patients with spinal muscular atrophy

Data from: Revised Airlie House consensus guidelines for design and implementation of ALS clinical trials

Objective: To revise the 1999 Airlie House consensus guidelines for the design and implementation of preclinical therapeutic studies and clinical trials in amyotrophic lateral sclerosis (ALS). Methods: A consensus committee comprising 140 key members of the international ALS community (ALS researchers, clinicians, patient representatives, research funding representatives, industry and regulatory agencies) addressed nine areas of need within ALS research: 1. Pre-clinical studies; 2. Biological and phenotypic heterogeneity; 3. Outcome measures; 4. Disease-modifying and symptomatic interventions; 5. Recruitment and retention; 6. Biomarkers; 7. Clinical trial phases; 8. Beyond traditional trial designs; and 9. Statistical considerations. Assigned to one of eight sections, committee members generated a draft set of recommendations based on a “background” of developing a (pre)clinical question and a “rationale” outlining the evidence and expert opinion. Following a two-day, face-to-face workshop at the Airlie House Conference Center, a modified Delphi process was used to develop draft consensus guidelines, which were subsequently reviewed and modified based on comments from the public. Statistical experts drafted a separate document of statistical considerations (section 9). Results: In this paper we summarize 112 recommendations and their associated backgrounds and rationales. The full list of recommendations on all main topics as well as on statistical considerations are available (Appendices e-3 and e-4). The authors prioritized 15 recommendations with the greatest potential to improve ALS clinical research. Conclusion: The revised Airlie House ALS Clinical Trials Consensus Guidelines should serve to improve clinical trial design and accelerate the development of effective treatments for patients with ALS

Appendix e-4 Full ALS Clinical Trial Guidelines

Appendix e-4 Full ALS Clinical Trial Guideline

Appendix e-5 Statistical Considerations

Appendix e-5 Statistical Consideration

Revised Airlie House consensus guidelines for design and implementation of ALS clinical trials.

ObjectiveTo revise the 1999 Airlie House consensus guidelines for the design and implementation of preclinical therapeutic studies and clinical trials in amyotrophic lateral sclerosis (ALS).MethodsA consensus committee comprising 140 key members of the international ALS community (ALS researchers, clinicians, patient representatives, research funding representatives, industry, and regulatory agencies) addressed 9 areas of need within ALS research: (1) preclinical studies; (2) biological and phenotypic heterogeneity; (3) outcome measures; (4) disease-modifying and symptomatic interventions; (5) recruitment and retention; (6) biomarkers; (7) clinical trial phases; (8) beyond traditional trial designs; and (9) statistical considerations. Assigned to 1 of 8 sections, committee members generated a draft set of guidelines based on a "background" of developing a (pre)clinical question and a "rationale" outlining the evidence and expert opinion. Following a 2-day, face-to-face workshop at the Airlie House Conference Center, a modified Delphi process was used to develop draft consensus research guidelines, which were subsequently reviewed and modified based on comments from the public. Statistical experts drafted a separate document of statistical considerations (section 9).ResultsIn this report, we summarize 112 guidelines and their associated backgrounds and rationales. The full list of guidelines, the statistical considerations, and a glossary of terms can be found in data available from Dryad (appendices e-3-e-5, doi.org/10.5061/dryad.32q9q5d). The authors prioritized 15 guidelines with the greatest potential to improve ALS clinical research.ConclusionThe revised Airlie House ALS Clinical Trials Consensus Guidelines should serve to improve clinical trial design and accelerate the development of effective treatments for patients with ALS

Appendix e-3 Glossary of Terms

Appendix e-3 Glossary of Term