AAV gene therapy for hereditary spastic paraplegia type 50: a phase 1 trial in a single patient.

There are more than 10,000 individual rare diseases and most are without therapy. Personalized genetic therapy represents one promising approach for their treatment. We present a road map for individualized treatment of an ultra-rare disease by establishing a gene replacement therapy developed for a single patient with hereditary spastic paraplegia type 50 (SPG50). Through a multicenter collaboration, an adeno-associated virus-based gene therapy product carrying the AP4M1 gene was created and successfully administered intrathecally to a 4-year-old patient within 3 years of diagnosis as part of a single-patient phase 1 trial. Primary endpoints were safety and tolerability, and secondary endpoints evaluated efficacy. At 12 months after dosing, the therapy was well tolerated. No serious adverse events were observed, with minor events, including transient neutropenia and Clostridioides difficile gastroenteritis, experienced but resolved. Preliminary efficacy measures suggest a stabilization of the disease course. Longer follow-up is needed to confirm the safety and provide additional insights on the efficacy of the therapy. Overall, this report supports the safety of gene therapy for SPG50 and provides insights into precision therapy development for rare diseases. Clinical trial registration: NCT06069687 .

Muscle MRI patterns for limb girdle muscle dystrophies: systematic review.

Limb girdle muscle dystrophies (LGMDs) are a group of inherited neuromuscular disorders comprising more than 20 genes. There have been increasing efforts to characterize this group with Muscle MRI. However, due to the complexity and similarities, the interpretation of the MRI patterns is usually done by experts in the field. Here, we proposed a step-by-step image interpretation of Muscle MRI in LGDM by evaluating the variability of muscle pattern involvement reported in the literature. A systematic review with an open start date to November 2022 was conducted to describe all LGMDs' muscle MRI patterns. Eighty-eight studies were included in the final review. Data were found to describe muscle MRI patterns for 15 out of 17 LGMDs types. Although the diagnosis of LGMDs is challenging despite the advanced genetic testing and other diagnostic modalities, muscle MRI is shown to help in the diagnosis of LGMDs. To further increase the yield for muscle MRI in the neuromuscular field, larger cohorts of patients need to be conducted.

Bending Drosophila larva using a microfluidic device enables imaging of its brain and nervous system at single neuronal resolution.

Single neuronal imaging of a fully intact Drosophila larva is a difficult challenge for neurosciences due to the robust digging/burrowing behaviour of the Drosophila larva and the lack of intact immobilization methods at single-neuron resolution. In this paper, for the first time, a simple microfluidic device to completely immobilize the brain and the CNS of a live, fully-functioning Drosophila larva for single neuronal imaging has been demonstrated. The design of the microfluidic device contains a unique clamping feature which pins and bends the body of the larva at 1/3rd of its length from the head. This simple twist combined with the pinning mechanism not only could stop the locomotion of the larva but also could immobilize the major movement of internal organs including the CNS. The results showed that the bent trap could keep the single neuron completely inside the field of view (FOV) (50 µm × 50 µm) over 10 min of confocal imaging. The range of motion in the x- and y-axis was approximately 8 µm and 2.5 µm, respectively. This corresponds to a range of 16% and 6% along the axis of the channel and across it compared to the size of the FOV (50 µm × 50 µm). The calcium activity of the single neurons in a 3rd instar GCaMP5 larva (Cha-Gal4/CyO; UAS-GCaMP5G/TM3) was measured while its mouth region was exposed to 20 mM sodium azide (NaN3) for 5 s. The results showed that the activity of the neurons has been statistically (p < 0.0005) increased (∼60%).

Scoliosis in Spinal Muscular Atrophy Type 1 in the Nusinersen Era.

Background and Objectives: The introduction of spinal muscular dystrophy (SMA)-modifying therapies, such as antisense oligonucleotide therapy, has changed the natural history of SMA. Most reports on treatment outcomes have focused on motor scores and respiratory function. The objective of this study is to document the development and progression of scoliosis in patients with SMA1 treated with nusinersen. Methods: A descriptive single-center study was conducted in patients with SMA1 who were treated with nusinersen before 6 months of age. Data were collected on patients who met criteria, including age at the first nusinersen dose, number of nusinersen doses, degree of scoliosis, respiratory parameters, feeding route, and motor scores at baseline and follow-up. The Children's Hospital of Philadelphia Infant Test of Neuromuscular Disorders (CHOP-INTEND) was subanalyzed using axial (AxS) and appendicular motor (ApS) scores to evaluate a possible correlation between scoliosis and axial muscle strength. Results: From our cohort, 31 percent (11/35) of patients had a diagnosis of SMA1. Sixty-three percent (7/11) met the inclusion criteria. All patients (7/7) showed initial improvement in their CHOP-INTEND scores in correlation with improvement on the ApS. Despite this, most patients did not show improvement in the AxS. Subsequently, all patients developed scoliosis in the first year of life with Cobb angles that ranged between 18° and 60°. Furthermore, total CHOP-INTEND scores had dropped in 2 patients alongside the development of a Cobb angle of >40°. Discussion: Despite the significant improvement in functional motor assessment in patients with SMA1, there is a progression of significant scoliosis despite treatment. Subsequently, lack or minimal improvement on the axial CHOP-INTEND scores may predict worsening on the total motor scores.

Diverse Oncogenic Fusions and Distinct Gene Expression Patterns Define the Genomic Landscape of Pediatric Papillary Thyroid Carcinoma.

Pediatric papillary thyroid carcinoma (PPTC) is clinically distinct from adult-onset disease. Although there are higher rates of metastasis and recurrence in PPTC, prognosis remains highly favorable. Molecular characterization of PPTC has been lacking. Historically, only 40% to 50% of childhood papillary thyroid carcinoma (PTC) were known to be driven by genomic variants common to adult PTC; oncogenic drivers in the remainder were unknown. This contrasts with approximately 90% of adult PTC driven by a discrete number of variants. In this study, 52 PPTCs underwent candidate gene testing, followed in a subset by whole-exome and transcriptome sequencing. Within these samples, candidate gene testing identified variants in 31 (60%) tumors, while exome and transcriptome sequencing identified oncogenic variants in 19 of 21 (90%) remaining tumors. The latter were enriched for oncogenic fusions, with 11 nonrecurrent fusion transcripts, including two previously undescribed fusions, STRN-RET and TG-PBF. Most fusions were associated with 3' receptor tyrosine kinase (RTK) moieties: RET, MET, ALK, and NTRK3. For advanced (distally metastatic) tumors, a driver variant was described in 91%. Gene expression analysis defined three clusters that demonstrated distinct expression of genes involved in thyroid differentiation and MAPK signaling. Among RET-CCDC6-driven tumors, gene expression in pediatric tumors was distinguishable from that in adults. Collectively, these results show that the genomic landscape of pediatric PTC is different from adult PTC. Moreover, they identify genomic drivers in 98% of PPTCs, predominantly oncogenic fusion transcripts involving RTKs, with a pronounced impact on gene expression. Notably, most advanced tumors were driven by a variant for which targeted systemic therapy exists. SIGNIFICANCE: This study highlights important distinctions between the genomes and transcriptomes of pediatric and adult papillary thyroid carcinoma, with implications for understanding the biology, diagnosis, and treatment of advanced disease in children.

A microfluidic microinjector for toxicological and developmental studies in Drosophila embryos.

Microinjection is an established and reliable method to deliver biological reagents such as transgenic constructs and drugs, to specific locations inside organisms such as the Drosophila embryo and C. elegans worm. In this paper, a simple compliant mechanism based PDMS-microinjection system has been demonstrated. Unlike conventional microinjectors, this unique system could allow one to precisely insert a long taper microneedle laterally and at various positions inside the length of the Drosophila embryo (up to 250 µm) with the resolution of 5 µm. Volumes as low as 30 pL with accuracy of ±10 pL were delivered inside the embryo via pressure pulses. The device has been used to study the effect of toxins on cardiogenesis in Drosophila embryos. Using this device, we demonstrate that the cardioblast (CB) migration velocity is modified in a dose sensitive manner to varying doses of injected sodium azide (NaN3) and, for the first time, quantify the effect of the toxin on heart assembly. Injection with 40 pL of NaN3 was shown to decrease CB migration velocity and filopodia number at concentrations above 10 mM, while embryos injected with the tracer Rhodamine B (0 mM NaN3) displayed no significant difference when compared to uninjected embryos. This device can be potentially used for other embryonic assays, which require accurate delivery of the reagents to a specific location within the embryo.