After nearly 40 years of development, oligonucleotide therapeutics are nearing meaningful clinical productivity. One of the key advantages of oligonucleotide drugs is that their delivery and potency are derived primarily from the chemical structure of the oligonucleotide whereas their target is defined by the base sequence. Thus, as oligonucleotides with a particular chemical design show appropriate distribution and safety profiles for clinical gene silencing in a particular tissue, this will open the door to the rapid development of additional drugs targeting other disease-associated genes in the same tissue. To achieve clinical productivity, the chemical architecture of the oligonucleotide needs to be optimized with a combination of sugar, backbone, nucleobase, and 3'- and 5'-terminal modifications. A portfolio of chemistries can be used to confer drug-like properties onto the oligonucleotide as a whole, with minor chemical changes often translating into major improvements in clinical efficacy. One outstanding challenge in oligonucleotide chemical development is the optimization of chemical architectures to ensure long-term safety. There are multiple designs that enable effective targeting of the liver, but a second challenge is to develop architectures that enable robust clinical efficacy in additional tissues.

The chemical evolution of oligonucleotide therapies of clinical utility.

The oligonucleotide therapeutics field has seen remarkable progress over the last few years with the approval of the first antisense drug and with promising developments in late stage clinical trials using siRNA or splice switching oligonucleotides. However, effective delivery of oligonucleotides to their intracellular sites of action remains a major issue. This review will describe the biological basis of oligonucleotide delivery including the nature of various tissue barriers and the mechanisms of cellular uptake and intracellular trafficking of oligonucleotides. It will then examine a variety of current approaches for enhancing the delivery of oligonucleotides. This includes molecular scale targeted ligand-oligonucleotide conjugates, lipid- and polymer-based nanoparticles, antibody conjugates and small molecules that improve oligonucleotide delivery. The merits and liabilities of these approaches will be discussed in the context of the underlying basic biology.

The delivery of therapeutic oligonucleotides.

Pharmacokinetics, biodistribution and cell uptake of antisense oligonucleotides.

CRISPR/Cas9-based therapeutics, especially those that can correct gene mutations via homology directed repair (HDR), have the potential to revolutionize the treatment of genetic diseases. However, HDR-based therapeutics are challenging to develop because they require simultaneous in vivo delivery of Cas9 protein, guide RNA and donor DNA. Here, we demonstrate that a delivery vehicle composed of gold nanoparticles conjugated to DNA and complexed with cationic endosomal disruptive polymers can deliver Cas9 ribonucleoprotein and donor DNA into a wide variety of cell types, and efficiently correct the DNA mutation that causes Duchenne muscular dystrophy in mice via local injection, with minimal off-target DNA damage.

Nanoparticle delivery of Cas9 ribonucleoprotein and donor DNA in vivo induces homology-directed DNA repair

Advances in biomaterials for drug delivery are enabling significant progress in biology and medicine. Multidisciplinary collaborations between physical scientists, engineers, biologists, and clinicians generate innovative strategies and materials to treat a range of diseases. Specifically, recent advances include major breakthroughs in materials for cancer immunotherapy, autoimmune diseases, and genome editing. Here, strategies for the design and implementation of biomaterials for drug delivery are reviewed. A brief history of the biomaterials field is first established, and then commentary on RNA delivery, responsive materials development, and immunomodulation are provided. Current challenges associated with these areas as well as opportunities to address long-standing problems in biology and medicine are discussed throughout.

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Advances in Biomaterials for Drug Delivery.

Objective
In prior open-label studies, eteplirsen, a phosphorodiamidate morpholino oligomer, enabled dystrophin production in Duchenne muscular dystrophy (DMD) with genetic mutations amenable to skipping exon 51. The present study used a double-blind placebo-controlled protocol to test eteplirsen's ability to induce dystrophin production and improve distance walked on the 6-minute walk test (6MWT).

Methods
DMD boys aged 7 to 13 years, with confirmed deletions correctable by skipping exon 51 and ability to walk 200 to 400 m on 6 MWT, were randomized to weekly intravenous infusions of 30 or 50 mg/kg/wk eteplirsen or placebo for 24 weeks (n = 4/group). Placebo patients switched to 30 or 50 mg/kg eteplirsen (n = 2/group) at week 25; treatment was open label thereafter. All patients had muscle biopsies at baseline and week 48. Efficacy included dystrophin-positive fibers and distance walked on the 6MWT.

Results
At week 24, the 30 mg/kg eteplirsen patients were biopsied, and percentage of dystrophin-positive fibers was increased to 23% of normal; no increases were detected in placebo-treated patients (p ≤ 0.002). Even greater increases occurred at week 48 (52% and 43% in the 30 and 50 mg/kg cohorts, respectively), suggesting that dystrophin increases with longer treatment. Restoration of functional dystrophin was confirmed by detection of sarcoglycans and neuronal nitric oxide synthase at the sarcolemma. Ambulation-evaluable eteplirsen-treated patients experienced a 67.3 m benefit compared to placebo/delayed patients (p ≤ 0.001).

Interpretation
Eteplirsen restored dystrophin in the 30 and 50 mg/kg/wk cohorts, and in subsequently treated, placebo-controlled subjects. Duration, more than dose, accounted for dystrophin production, also resulting in ambulation stability. No severe adverse events were encountered. Ann Neurol 2013;74:637–647

https://onlinelibrary.wiley.com/doi/epdf/10.1002/ana.23982

Eteplirsen for the treatment of Duchenne muscular dystrophy

➤ Limb-length discrepancy is a common condition found in the general population and elderly individuals with osteoarthritis.

➤ All aspects of total hip arthroplasty are interconnected, and making adjustments in one can affect several others.

➤ Intraoperative imaging with trial or final components in place is helpful to rule out excessive limb-length discrepancy and to confirm proper component sizing and position in difficult primary total hip arthroplasty or revision situations.

Limb-Length Discrepancy After Hip Arthroplasty

BACKGROUND Duchenne muscular dystrophy (DMD) possesses many potential challenges for anesthetic care. Invasive and noninvasive procedures with corresponding sedation or general anesthesia are frequent and necessary for affected patients. There remains a need for a better agent or agents for procedural sedation in patients with comorbid diseases. This study prospectively evaluated a combination of ketamine with two different doses of dexmedetomidine for sedation during muscle biopsy in patients with DMD. METHODS Dexmedetomidine 1.0 or 0.5 μg·kg(-1) was administered as a loading dose over 3 min followed by a continuous infusion of 1.0 or 0.5 μg·kg·h(-1). Ketamine (1 mg·kg(-1)) was administered along with the dexmedetomidine loading dose. As the procedure commenced, additional doses of ketamine (0.5 mg·kg(-1)) were administered as needed. Sedation scores, hemodynamic data, operative times, and recovery times were recorded. RESULTS The study cohort included a total of 53 bicep, deltoid, or anterior tibialis muscle biopsies in 19 boys including 24 in the dexmedetomidine 1.0 μg·kg(-1) group and 29 in the dexmedetomidine 0.5 μg·kg(-1) group. Mean age and weight were 9.7 ± 1.4 years and 33.3 ± 7.7 kg in the dexmedetomidine 1.0 μg·kg(-1) group and 8.8 ± 1.8 years and 30.2 ± 10.8 kg in the dexmedetomidine 0.5 μg·kg(-1) group. No significant changes in blood pressure were noted. A decrease in heart rate (HR) occurred after the loading dose of dexmedetomidine in both groups. The HR was significantly lower in the dexmedetomidine 1.0 μg·kg(-1) group compared with the dexmedetomidine 0.5 μg·kg(-1) group. Total recovery time to discharge was significantly shorter in the dexmedetomidine 0.5 μg·kg(-1) group than the dexmedetomidine 1.0 μg·kg(-1) group (146 ± 65 vs 174 ± 58 min; P = 0.03), although the total ketamine dose was significantly greater in the dexmedetomidine 0.5 μg·kg(-1) group (3.7 ± 1.0 vs 2.0 ± 0.5 mg·kg(-1); P < 0.01). There were no episodes of apnea or hypoventilation; however, a jaw thrust was needed in one patient in the dexmedetomidine 1.0 μg·kg(-1) group. CONCLUSION The combination of dexmedetomidine and ketamine is safe and effective for moderately painful procedures with limited respiratory and cardiovascular effects in a high-risk patient population. Dexmedetomidine 0.5 μg·kg(-1) as a loading dose with ketamine followed by a continuous infusion of dexmedetomidine at 0.5 μg·kg(-1) ·h(-1) achieved an adequate sedation level with shorter total recovery times in the perioperative unit compared with a higher dose regimen of dexmedetomidine (1.0 μg·kg(-1) loading dose followed by an infusion at 1.0 μg·kg(-1) ·h(-1)).

Dexmedetomidine and ketamine sedation for muscle biopsies in patients with Duchenne muscular dystrophy

US readily demonstrates cartilaginous structures, and static sonography has shown potential in evaluating clubfoot deformity. To investigate the potential of dynamic sonography in the evaluation of the congenital clubfoot. Sonography was used for static and dynamic stress evaluation of 13 clubfeet and 35 normal feet in 24 patients (ages 0–32 weeks). Dynamic foot sonography was performed using a single-operator bimanual scanning technique. The examination involved coronal oblique evaluation of the medial malleolar–navicular (MMN) distance and the calcaneocuboid relationship, sagittal evaluation of the talonavicular relationship, and transverse evaluation of navicular subluxation, rotation, and deformation. Dynamic abduction/adduction stress maneuvers were performed, measured by the MMN. The clubfoot “gristle” is a consistent, measurable soft-tissue landmark in clubfeet, connecting the medial malleolus to the medial navicular and talus. Mean MMN distances in clubfeet in the neutral position and abduction were significantly different from these distances in the normal paired foot (differences of 8.7 mm neutral position and 7.94 mm abduction), as compared to bilateral normal feet (differences of 0.98 mm neutral position and 1.43 mm abduction). Navicular subluxation showed good correlation between highly deformed and subluxated navicular bones and a tight medial clubfoot complex. Focused dynamic foot sonography is useful in providing a specific and detailed functional preoperative and/or postoperative assessment of the congenital clubfoot.

John R. Kean

Papers

Eteplirsen for the treatment of Duchenne muscular dystrophy

Limb-Length Discrepancy After Hip Arthroplasty

Dexmedetomidine and ketamine sedation for muscle biopsies in patients with Duchenne muscular dystrophy

Focused dynamic sonographic examination of the congenital clubfoot.

Eteplirsen for the treatment of duchenne muscular dystrophy. Ann Neurol