Designing clinical trials for rare diseases: unique challenges and opportunities

Orphan drug development is a rapidly expanding field. Nevertheless, clinical trials for rare diseases can present inherent challenges. Optimal study design and partnerships between academia and industry are therefore required for the successful development, delivery and clinical approval of effective therapies in this group of disorders

FCI: an R-based algorithm for evaluating uncertainty of absolute real-time PCR quantification

Background: FCI is an R code for analyzing data from real-time PCR experiments. This algorithm estimates standard curve features as well as nucleic acid concentrations and confidence intervals according to Fieller's theorem. Results: In order to describe the features of FCI four situations were selected from real data collected during an international external quality assessment program for quantitative assays based on real-time PCR. The code generates a diagnostic figure suitable for assessing the quality of the quantification process. Conclusion: We have provided a freeware programme using this algorithm specifically designed to increase the information content of the real-time PCR assay. \ua9 2008 Verderio et al; licensee BioMed Central Ltd

Phenotype and genotype of 197 British patients with McArdle disease: An observational single-centre study

McArdle disease is caused by recessive mutations in PYGM gene. The condition is considered to cause a “pure” muscle phenotype with symptoms including exercise intolerance, inability to perform isometric activities, contracture, and acute rhabdomyolysis leading to acute renal failure. This is a retrospective observational study aiming to describe phenotypic and genotypic features of a large cohort of patients with McArdle disease between 2011 and 2019. Data relating to genotype and phenotype, including frequency of rhabdomyolysis, fixed muscle weakness, gout and comorbidities, inclusive of retinal disease (pattern retinal dystrophy) and thyroid disease, were collected. Data from 197 patients are presented. Seven previously unpublished PYGM mutations are described. Exercise intolerance (100%) and episodic rhabdomyolysis (75.6%) were the most common symptoms. Fixed muscle weakness was present in 82 (41.6%) subjects. Unexpectedly, ptosis was observed in 28 patients (14.2%). Hyperuricaemia was a common finding present in 88 subjects (44.7%), complicated by gout in 25% of cases. Thyroid dysfunction was described in 30 subjects (15.2%), and in 3 cases, papillary thyroid cancer was observed. Pattern retinal dystrophy was detected in 15 out of the 41 subjects that underwent an ophthalmic assessment (36.6%). In addition to fixed muscle weakness, ptosis was a relatively common finding. Surprisingly, dysfunction of thyroid and retinal abnormalities were relatively frequent comorbidities. Further studies are needed to better clarify this association, although our finding may have important implication for patient management

Mitochondrial Strokes: Diagnostic Challenges and Chameleons

Mitochondrial stroke-like episodes (SLEs) are a hallmark of mitochondrial encephalomyopathy, lactic acidosis, and stroke-like episodes (MELAS). They should be suspected in anyone with an acute/subacute onset of focal neurological symptoms at any age and are usually driven by seizures. Suggestive features of an underlying mitochondrial pathology include evolving MRI lesions, often originating within the posterior brain regions, the presence of multisystemic involvement, including diabetes, deafness, or cardiomyopathy, and a positive family history. The diagnosis of MELAS has important implications for those affected and their relatives, given it enables early initiation of appropriate treatment and genetic counselling. However, the diagnosis is frequently challenging, particularly during the acute phase of an event. We describe four cases of mitochondrial strokes to highlight the considerable overlap that exists with other neurological disorders, including viral and autoimmune encephalitis, ischemic stroke, and central nervous system (CNS) vasculitis, and discuss the clinical, laboratory, and imaging features that can help distinguish MELAS from these differential diagnoses

Cortical visuomotor interactions in Freezing of Gait: A TMS approach

OBJECTIVES: Altered cortical visuomotor integration has been involved in the pathophysiology of freezing of gait (FoG) in parkinsonism. The aim of this study was to assess the connections between the primary visual (V1) and motor (M1) areas with a paired-pulse, twin-coil transcranial magnetic stimulation (TMS) technique in patients with FoG. METHODS: Twelve Parkinson's disease (PD) patients suffering from levodopa-responsive-FoG (off-FoG) were compared with 12 PD patients without FoG and 12 healthy subjects of similar age/sex. In the "off" condition, visuomotor connections (VMCs) were assessed bilaterally. A conditioning stimulus over the V1 phosphene hotspot was followed at interstimulus intervals (ISIs) of 18 and 40ms by a test stimulus over M1, to elicit motor evoked potentials (MEPs) in the contralateral first dorsal interosseous muscle. RESULTS: Significant (P<0.01), bilateral effects due to VMCs were detected in all three groups, consisting of a MEP suppression at ISI 18 and 40ms. However, in PD patients with FoG, the MEP suppression was significantly (P<0.05) enhanced, both at ISI 18-40ms, in comparison with the other two groups. The phenomenon was limited to the right hemisphere. CONCLUSIONS: PD patients with FoG showed an excessive inhibitory response of the right M1 to inputs travelling from V1 at given ISIs. Right-sided alterations of the cortical visuomotor integration may contribute to the pathophysiology of FoG

Insights on the bacterial composition of Parmigiano Reggiano Natural Whey Starter by a culture-dependent and 16S rRNA metabarcoding portrait

: Natural whey starters (NWS) are undefined bacterial communities produced daily from whey of the previous cheese-making round, by application of high temperature. As a result, in any dairy plant, NWS are continuously evolving, undefined mixtures of several strains and/or species of lactic acid bacteria, whose composition and performance strongly depend on the selective pressure acting during incubation. While NWS is critical to assure consistency to cheese-making process, little is known about the composition, functional features, and plant-to-plant fluctuations. Here, we integrated 16S rRNA metabarcoding and culture-dependent methods to profile bacterial communities of 10 NWS sampled in the production area of Parmigiano Reggiano cheese. 16S rRNA metabarcoding analysis revealed two main NWS community types, namely NWS type-H and NWS type-D. Lactobacillus helveticus was more abundant in NWS type-H, whilst Lactobacillus delbrueckii/St. thermophilus in NWS type-D, respectively. Based on the prediction of metagenome functions, NWS type-H samples were enriched in functional pathways related to galactose catabolism and purine metabolism, while NWS type-D in pathways related to aromatic and branched chain amino acid biosynthesis, which are flavor compound precursors. Culture-dependent approaches revealed low cultivability of individual colonies as axenic cultures and high genetic diversity in the pool of cultivable survivors. Co-culturing experiments showed that fermentative performance decreases by reducing the bacterial complexity of inoculum, suggesting that biotic interactions and cross-feeding relationships could take place in NWS communities, assuring phenotypic robustness. Even though our data cannot directly predict these ecological interactions, this study provides the basis for experiments targeted at understanding how selective regime affects composition, bacterial interaction, and fermentative performance in NWS

Tracing the identity of Parmigiano Reggiano “Prodotto di Montagna - Progetto Territorio” cheese using NMR spectroscopy and multivariate data analysis

Background Nuclear magnetic resonance (NMR) spectroscopy is one of the well-established tools for food metabolomic analysis, as it proved to be very effective in authenticity and quality control of dairy products, as well as to follow product evolution during processing and storage. The analytical assessment of the EU mountain denomination label, specifically for Parmigiano Reggiano "Prodotto di Montagna - Progetto Territorio" (Mountain-CQ) cheese, has received limited attention. Although it was established in 2012 the EU mountain denomination label has not been much studied from an analytical point of view. Nonetheless, tracing a specific profile for the mountain products is essential to support the value chain of this specialty. Results The aim of the study was to produce an identity profile for Parmigiano Reggiano “Prodotto di Montagna - Progetto Territorio” (Mountain-CQ) cheese, and to differentiate it from Parmigiano Reggiano PDO samples (conventional-PDO) using 1H NMR spectroscopy coupled with multivariate data analysis. Three different approaches were applied and compared. First, the spectra-as-such were analysed after proper preprocessing. For the other two approaches, Multivariate Curve Resolution-Alternating Least Squares (MCR-ALS) was used for signals resolution and features extraction, either individually on manually-defined spectral intervals or by reapplying MCR-ALS on the whole spectra with selectivity constraints using the reconstructed “pure profiles” as initial estimates and targets. All approaches provided comparable information regarding the samples’ distribution, as in all three cases the separation between the two product categories conventional-PDO and Mountain-CQ could be highlighted. Moreover, a novel MATLAB toolbox for features extraction via MCR-ALS was developed and used in synergy with the Chenomx library, allowing for a putative identification of the selected features. Significance A first identity profile for Parmigiano Reggiano “Prodotto di Montagna - Progetto Territorio” obtained by interpreting the metabolites signals in NMR spectroscopy was obtained. Our workflow and toolbox for generating the features dataset allows a more straightforward interpretation of the results, to overcome the limitations due to dimensionality and to peaks overlapping, but also to include the signals assignment and matching since the early stages of the data processing and analysis

Factors Affecting the Clinical Measurement of Visuo-Spatial Neglect

The present study examined a battery of tests to evaluate unilateral spatial neglect; the tests included different tasks involving several modalities of spatial exploration mapping perceptual, motor, attentional and personal or extrapersonal space dimensions. The subjects, 121 right-brain-damaged patients with unilateral neglect, were studied in seven laboratories in four European countries. Relationships among the various tests were examined by correlations, a cluster analysis and by an analysis of individual cases. Different sensitivity was found among various tests for detecting neglect performances. Both the cluster analysis and the single case analysis clearly showed a segregation between personal and extrapersonal neglect. Analysis of the large cluster, including a variety of tests of extra personal neglect, together with the study of single cases, suggests the possibility of differentiating the various manifestations of spatial neglect which can be interpreted on the basis of the descriptions of other individual cases previously reported in the literature. Finally, the present study indicated the relative stability of neglect following the acute phase and its independence from age

First presentation of LPIN1 acute rhabdomyolysis in adolescence and adulthood

LPIN1 mutations are a known common cause of autosomal recessive, recurrent and life-threatening acute rhabdomyolysis of childhood-onset. The first episode of rhabdomyolysis usually happens in nearly all cases before the age of 5 and death is observed in 1/3 of patients. Here we present two cases of acute rhabdomyolysis with a milder phenotype caused by LPIN1 mutation presenting in adolescence (11 years old) and adulthood (40 years old) after Parvovirus infection and metabolic stress, respectively. In our opinion, the mutation types, epigenetic factors, the environment exposition to triggers or the existence of proteins with a similar structure of LPIN1, may have a role in modulating the onset of rhabdomyolysis. LPIN1 should be included on a panel of genes analysed in the investigation of adult individuals with rhabdomyolysis. Metabolic and viral stressors should be included in the list of possible rhabdomyolysis precipitant