Translational and Regulatory Challenges for Exon Skipping Therapies

Several translational challenges are currently impeding the therapeutic development of antisense-mediated exon skipping approaches for rare diseases. Some of these are inherent to developing therapies for rare diseases, such as small patient numbers and limited information on natural history and interpretation of appropriate clinical outcome measures. Others are inherent to the antisense oligonucleotide (AON)-mediated exon skipping approach, which employs small modified DNA or RNA molecules to manipulate the splicing process. This is a new approach and only limited information is available on long-term safety and toxicity for most AON chemistries. Furthermore, AONs often act in a mutation-specific manner, in which case multiple AONs have to be developed for a single disease. A workshop focusing on preclinical development, trial design, outcome measures, and different forms of marketing authorization was organized by the regulatory models and biochemical outcome measures working groups of Cooperation of Science and Technology Action: "Networking towards clinical application of antisense-mediated exon skipping for rare diseases." The workshop included participants from patient organizations, academia, and members of staff from the European Medicine Agency and Medicine Evaluation Board (the Netherlands). This statement article contains the key outcomes of this meeting.status: publishe

Antisense PMO Found in Dystrophic Dog Model Was Effective in Cells from Exon 7-Deleted DMD Patient

BACKGROUND: Antisense oligonucleotide-induced exon skipping is a promising approach for treatment of Duchenne muscular dystrophy (DMD). We have systemically administered an antisense phosphorodiamidate morpholino oligomer (PMO) targeting dystrophin exons 6 and 8 to a dog with canine X-linked muscular dystrophy in Japan (CXMD(J)) lacking exon 7 and achieved recovery of dystrophin in skeletal muscle. To date, however, antisense chemical compounds used in DMD animal models have not been directly applied to a DMD patient having the same type of exon deletion. We recently identified a DMD patient with an exon 7 deletion and tried direct translation of the antisense PMO used in dog models to the DMD patient's cells. METHODOLOGY/PRINCIPAL FINDINGS: We converted fibroblasts of CXMD(J) and the DMD patient to myotubes by FACS-aided MyoD transduction. Antisense PMOs targeting identical regions of dog and human dystrophin exons 6 and 8 were designed. These antisense PMOs were mixed and administered as a cocktail to either dog or human cells in vitro. In the CXMD(J) and human DMD cells, we observed a similar efficacy of skipping of exons 6 and 8 and a similar extent of dystrophin protein recovery. The accompanying skipping of exon 9, which did not alter the reading frame, was different between cells of these two species. CONCLUSION/SIGNIFICANCE: Antisense PMOs, the effectiveness of which has been demonstrated in a dog model, achieved multi-exon skipping of dystrophin gene on the FACS-aided MyoD-transduced fibroblasts from an exon 7-deleted DMD patient, suggesting the feasibility of systemic multi-exon skipping in humans

A Hybrid Model for Dynamic Simulation of Custom Software Projects in a Multiproject Environment

This paper describes SimHiProS, a hybrid simulation model of software production. The goal is to gain insight on the dynamics induced by resource sharing in multiproject management. In order to achieve it the hierarchy of decisions in a multiproject organization is modeled and some resource allocation methods based on algorithms from the OR/AI domain are used. Other critical issues such as the hybrid nature of software production and the effects of measurement and control are also incorporated in the model. Some first results are presented.Ministerio de Ciencia e Innovación TIN2004-06689-C03-03Ministerio de Ciencia e Innovación TIN2007-67843-C06-0

Thermodynamic analysis of the Quantum Critical behavior of Ce-lattice compounds

A systematic analysis of low temperature magnetic phase diagrams of Ce compounds is performed in order to recognize the thermodynamic conditions to be fulfilled by those systems to reach a quantum critical regime and, alternatively, to identify other kinds of low temperature behaviors. Based on specific heat ($C_m$) and entropy ($S_m$) results, three different types of phase diagrams are recognized: i) with the entropy involved into the ordered phase ($S_{MO}$) decreasing proportionally to the ordering temperature ($T_{MO}$), ii) those showing a transference of degrees of freedom from the ordered phase to a non-magnetic component, with their $C_m(T_{MO})$ jump ($\Delta C_m$) vanishing at finite temperature, and iii) those ending in a critical point at finite temperature because their $\Delta C_m$ do not decrease with $T_{MO}$ producing an entropy accumulation at low temperature. Only those systems belonging to the first case, i.e. with $S_{MO}\to 0$ as $T_{MO}\to 0$, can be regarded as candidates for quantum critical behavior. Their magnetic phase boundaries deviate from the classical negative curvature below $T\approx 2.5$\,K, denouncing frequent misleading extrapolations down to T=0. Different characteristic concentrations are recognized and analyzed for Ce-ligand alloyed systems. Particularly, a pre-critical region is identified, where the nature of the magnetic transition undergoes significant modifications, with its $\partial C_m/\partial T$ discontinuity strongly affected by magnetic field and showing an increasing remnant entropy at $T\to 0$. Physical constraints arising from the third law at $T\to 0$ are discussed and recognized from experimental results

The impact of Patient and Public Involvement in the SlowMo study: Reflections on peer innovation

BACKGROUND: The SlowMo study demonstrated the effects of SlowMo, an eight-session digitally supported reasoning intervention, on paranoia in a large-scale randomized-controlled trial with 362 participants with schizophrenia-spectrum psychosis. AIM: The current evaluation aimed to investigate the impact of Patient and Public Involvement (PPI) in the SlowMo study. METHOD: PPI members were six women and three men from Sussex, Oxford and London with experience of using mental health services for psychosis. They received training and met at least 3-monthly throughout the project. The impact of PPI was captured quantitatively and qualitatively through (i) a PPI log of recommendations and implementation; (ii) written subjective experiences of PPI members; (iii) meeting minutes; and (iv) outputs produced. RESULTS: The PPI log revealed 107 recommendations arising from PPI meetings, of which 87 (81%) were implemented. Implementation was greater for recruitment-, data collection- and organization-related actions than for dissemination and emergent innovations. Qualitative feedback revealed impacts on study recruitment, data collection, PPI participants' confidence, knowledge, career aspirations and society more widely. Outputs produced included a film about psychosis that aired on BBC primetime television, novel webpages and journal articles. Barriers to PPI impact included geography, travel, funding, co-ordination and well-being. DISCUSSION: A future challenge for PPI impact will be the extent to which peer innovation (innovative PPI-led ideas) can be supported within research study delivery. PATIENT AND PUBLIC CONTRIBUTION: Planned Patient and Public Contribution in SlowMo comprised consultation and collaboration in (i) design, (ii) recruitment, (iii) qualitative interviews and analysis of service users' experiences of SlowMo therapy and (iv) dissemination

How managers can build trust in strategic alliances: a meta-analysis on the central trust-building mechanisms

Trust is an important driver of superior alliance performance. Alliance managers are influential in this regard because trust requires active involvement, commitment and the dedicated support of the key actors involved in the strategic alliance. Despite the importance of trust for explaining alliance performance, little effort has been made to systematically investigate the mechanisms that managers can use to purposefully create trust in strategic alliances. We use Parkhe’s (1998b) theoretical framework to derive nine hypotheses that distinguish between process-based, characteristic-based and institutional-based trust-building mechanisms. Our meta-analysis of 64 empirical studies shows that trust is strongly related to alliance performance. Process-based mechanisms are more important for building trust than characteristic- and institutional-based mechanisms. The effects of prior ties and asset specificity are not as strong as expected and the impact of safeguards on trust is not well understood. Overall, theoretical trust research has outpaced empirical research by far and promising opportunities for future empirical research exist

A proposal for multi-tens of GW fully coherent femtosecond soft X-ray lasers

X-ray free-electron lasers1,2 delivering up to 131013 coherent photons in femtosecond pulses are bringing about a revolution in X-ray science3?5. However, some plasma-based soft X-ray lasers6 are attractive because they spontaneously emit an even higher number of photons (131015), but these are emitted in incoherent and long (hundreds of picoseconds) pulses7 as a consequence of the amplification of stochastic incoherent self-emission. Previous experimental attempts to seed such amplifiers with coherent femtosecond soft X-rays resulted in as yet unexplained weak amplification of the seed and strong amplification of incoherent spontaneous emission8. Using a time-dependent Maxwell?Bloch model describing the amplification of both coherent and incoherent soft X-rays in plasma, we explain the observed inefficiency and propose a new amplification scheme based on the seeding of stretched high harmonics using a transposition of chirped pulse amplification to soft X-rays. This scheme is able to deliver 531014 fully coherent soft X-ray photons in 200 fs pulses and with a peak power of 20 GW

Development of Exon Skipping Therapies for Duchenne Muscular Dystrophy: A Critical Review and a Perspective on the Outstanding Issues

Functional Genomics of Muscle, Nerve and Brain Disorder

Dynamics of Co-Transcriptional Pre-mRNA Folding Influences the Induction of Dystrophin Exon Skipping by Antisense Oligonucleotides

Antisense oligonucleotides (AONs) mediated exon skipping offers potential therapy for Duchenne muscular dystrophy. However, the identification of effective AON target sites remains unsatisfactory for lack of a precise method to predict their binding accessibility. This study demonstrates the importance of co-transcriptional pre-mRNA folding in determining the accessibility of AON target sites for AON induction of selective exon skipping in DMD. Because transcription and splicing occur in tandem, AONs must bind to their target sites before splicing factors. Furthermore, co-transcriptional pre-mRNA folding forms transient secondary structures, which redistributes accessible binding sites. In our analysis, to approximate transcription elongation, a “window of analysis” that included the entire targeted exon was shifted one nucleotide at a time along the pre-mRNA. Possible co-transcriptional secondary structures were predicted using the sequence in each step of transcriptional analysis. A nucleotide was considered “engaged” if it formed a complementary base pairing in all predicted secondary structures of a particular step. Correlation of frequency and localisation of engaged nucleotides in AON target sites accounted for the performance (efficacy and efficiency) of 94% of 176 previously reported AONs. Four novel insights are inferred: (1) the lowest frequencies of engaged nucleotides are associated with the most efficient AONs; (2) engaged nucleotides at 3′ or 5′ ends of the target site attenuate AON performance more than at other sites; (3) the performance of longer AONs is less attenuated by engaged nucleotides at 3′ or 5′ ends of the target site compared to shorter AONs; (4) engaged nucleotides at 3′ end of a short target site attenuates AON efficiency more than at 5′ end