What Can We Learn From Clinical Trials of Exon Skipping for DMD?

On 20 September 2013, GlaxoSmithKline (GSK) and Prosensa announced that GSK’s Phase III clinical trial (NCT01254019) of Drisapersen, an exon skipping drug for Duchenne muscular dystrophy (DMD), failed to meet the primary endpoint of a statistically significant improvement in the 6 Minute Walking Distance Test (6MWT) compared to placebo.1 On 12 November 2013, Sarepta Therapeutics announced that the US Food and Drug Administration (FDA) has considered its application for accelerated approval of Eteplirsen as a DMD drug to be premature.2 The news came as a great disappointment to the scientific community and more specifically to the families and foundations that had followed the trail of research articles and press announcements. Moreover, the clinical trial results and the FDA’s view of the relationship between the dystrophin biomarker and functional endpoint of 6MWT in clinic will have a far reaching impact beyond exon skipping therapy in DMD. Currently, the most promising therapies for DMD are arguably gene therapy and exon skipping, both aiming to restore the expression of dystrophin. For exon skipping, the general strategy of restoring expression of the mutated dystrophin gene by excluding frame-disrupting mutations was vindicated by early experiments in dystrophic animal models.3–6 This principle has been substantiated for DMD by clinical trials over the last 7 years with two chemistries, the 2’O-methyl phosphorothioate backbone (2OMePS, named PRO051/ Drisapersen initiated by Prosensa/GSK) and the morpholino backbone (PMO, named Eteplirsen initiated by AVI Biopharma, now Sarepta Therapeutics).7–10 Both drugs target dystrophin exon 51 and both elicited the expected skipping of exon 51 and production of dystrophin protein following intramuscular injections. In a subsequent open-label, dose-escalation systemic study, five weekly subcutaneous injections of Prosensa/ GSK’s PRO051 at 0.5, 2, 4, or 6 mg/kg induced skipping of exon 51 accompanied by low levels of dystrophin in 12 DMD boys. But, importantly, this data lacked pretreatment controls.10 This lack of pretreatment controls in combination with the use of the highly sensitive dystrophin antibody MANDYS106 led to reports of expansion of dystrophin positive fibre counts to up to 100%. The follow-up extension for 12-weeks at 6 mg/kg of the PRO051 reported stabilisation of motor function in the boys.10 However, the study included several boys below 7 years of age where natural history would predict improved motor function within the experimental period. A subsequent Phase IIb placebo-controlled 6 mg/ kg/week study (NCT01153932) of Drisapersen on 53 DMD patients again reported a significant benefit in 6MWT in the treatment over the placebo group. Clinical benefits were maintained, but with reduced significance, after 48 weeks of treatment. However, no muscle biopsy results have yet been reported.11 This was followed by the phase III trial, with 186 patients, that failed to show statistically significant improvements in the primary outcome measure of the 6MWT. The difference in conclusion between the Phase III and earlier studies with the exact same treatment regime is therefore attributed almost solely to the high variability of the 6MWT endpoint measurement within the time-frame of the subject population and difference in sample sizes. What have we learnt from the Prosensa/GSK trials? According to the comment from the FDA in response to Sarepta’s application for Eteplirsen targeting the same dystrophin exon 51, the failure of the Drisapersen trial indicates a “disconnect between increased expression of dystrophin and clinical efficacy.” This criticism has serious implications far beyond exon skipping in DMD raising questions about our assessment of all experimental therapies that aim to restore or produce functional dystrophins in DMD, including gene replacement therapies that deliver a known functional gene product as a drug. So, is there clear evidence indicating a disconnect between the levels of dystrophin expression and clinical efficacy? Fortunately, the answer is no. Results, both from animal models6 and clinical studies on Becker muscular dystrophy (BMD) patients, all point to a positive connection. Much of the confusion about levels of dystrophin expression come from over emphasizing assessment of dystrophin expression solely by immunohistochemistry (IHC) without pretreatment control biopsies for each patient and the use of highly sensitive antibody MANDYS106. Prosensa reported that up to 100% of fibers were dystrophin positive in the muscle samples of some Drisapersen-treated patients in its phase II study.10 IHC is critical to provide us with a rough estimate of the levels of protein expression, the proportion of cells expressing the protein, and especially patterns of distribution. However, one must be aware that judgment of positivity of dystrophin What Can We Learn From Clinical Trials of Exon Skipping for DMD?