Frequency, mutual exclusivity and clinical associations of myositis autoantibodies in a combined European cohort of idiopathic inflammatory myopathy patients.

OBJECTIVES: To determine prevalence and co-existence of myositis specific autoantibodies (MSAs) and myositis associated autoantibodies (MAAs) and associated clinical characteristics in a large cohort of idiopathic inflammatory myopathy (IIM) patients. METHODS: Adult patients with confirmed IIM recruited to the EuroMyositis registry (n = 1637) from four centres were investigated for the presence of MSAs/MAAs by radiolabelled-immunoprecipitation, with confirmation of anti-MDA5 and anti-NXP2 by ELISA. Clinical associations for each autoantibody were calculated for 1483 patients with a single or no known autoantibody by global linear regression modelling. RESULTS: MSAs/MAAs were found in 61.5% of patients, with 84.7% of autoantibody positive patients having a sole specificity, and only three cases (0.2%) having more than one MSA. The most frequently detected autoantibody was anti-Jo-1 (18.7%), with a further 21 specificities each found in 0.2-7.9% of patients. Autoantibodies to Mi-2, SAE, TIF1, NXP2, MDA5, PMScl and the non-Jo-1 tRNA-synthetases were strongly associated (p < 0.001) with cutaneous involvement. Anti-TIF1 and anti-Mi-2 positive patients had an increased risk of malignancy (OR 4.67 and 2.50 respectively), and anti-SRP patients had a greater likelihood of cardiac involvement (OR 4.15). Interstitial lung disease was strongly associated with the anti-tRNA synthetases, anti-MDA5, and anti-U1RNP/Sm. Overlap disease was strongly associated with anti-PMScl, anti-Ku, anti-U1RNP/Sm and anti-Ro60. Absence of MSA/MAA was negatively associated with extra-muscular manifestations. CONCLUSIONS: Myositis autoantibodies are present in the majority of patients with IIM and identify distinct clinical subsets. Furthermore, MSAs are nearly always mutually exclusive endorsing their credentials as valuable disease biomarkers.

Plug-and-Play Benzylisoquinoline Alkaloid Biosynthetic Gene Discovery in Engineered Yeast.

Benzylisoquinoline alkaloid (BIA) metabolism has been the focus of a considerable research effort over the past half-century, primarily because of the pharmaceutical importance of several compounds produced by opium poppy (Papaver somniferum). Advancements in genomics technologies have substantially accelerated the rate of gene discovery over the past decade, such that most biosynthetic enzymes involved in the formation of the major alkaloids of opium poppy have now been isolated and partially characterized. Not unexpectedly, the availability of all perceived biosynthetic genes has facilitated the reconstitution of several BIA pathways in microbial hosts, including yeast (Saccharomyces cerevisiae). Product yields are currently insufficient to consider the commercial production of high-value BIAs, such as morphine. However, the rudimentary success demonstrated by the uncomplicated and routine assembly of a multitude of characterized BIA biosynthetic genes provides a valuable gene discovery tool for the rapid functional identification of the plethora of gene candidates available through increasingly accessible genomic, transcriptomic, and proteomic databases. BIA biosynthetic gene discovery represents a substantial research opportunity largely owing to the wealth of existing enzyme data mostly obtained from a single plant species. Functionally novel enzymes and variants with potential metabolic engineering applications can be considered the primary targets. Selection of candidates from sequence repositories is facilitated by the monophyletic relationship among biosynthetic genes belonging to a wide range of enzyme families, such as the numerous cytochromes P450 and AdoMet-dependent O- and N-methyltransferases that operate in BIA metabolism. We describe methods for the rapid functional screening of uncharacterized gene candidates encoding potential BIA biosynthetic enzymes using yeast strains engineered to perform selected metabolic conversions. As an initial screening tool, the approach is superior to the in vitro characterization of recombinant enzyme candidates, and provides a standardized functional genomics opportunity for otherwise recalcitrant exotic plant species.

Higher proportion of fast-twitch (type II) muscle fibres in idiopathic inflammatory myopathies - evident in chronic but not in untreated newly diagnosed patients.

OBJECTIVE: Polymyositis and dermatomyositis are idiopathic, inflammatory myopathies characterized by proximal muscle fatigue. Conventional immunosuppressive treatment gives a variable response. Biopsies from chronic patients display a low proportion type I and a high proportion of type II muscle fibres. This raised a suspicion that the low proportion of type I fibres might play a role in the muscle fatigue. AIM: To investigate whether the muscle fibre attributes evident in chronic myositis are characteristic for the polymyositis and dermatomyosistis diseases themselves. METHODS: Muscle biopsies were obtained from thigh muscle from untreated patients (n = 18), treated responders (n = 14) and non-responders (n = 6) and from healthy controls (n = 11), respectively. For clinical evaluations, creatine kinase, functional index of myositis and cumulative dose of cortisone were established. RESULTS: Chronic patients had a lower proportion of type I fibres and a higher proportion of type II fibres compared to untreated myositis patients and healthy controls. Fibre cross-sectional area (CSA) did not differ between patients and healthy individuals but all women had a 20% smaller type II fibre CSA compared to men. CONCLUSIONS: Untreated polymyositis and dermatomyositis patients and healthy controls have a different fibre type composition than chronic polymyositis and dermatomyositis patients. Fibre CSA did not differ between healthy controls or any of the patient groups. A low proportion of oxidative muscle fibres can therefore be excluded as a contributing factor causing muscle fatigue at disease onset and the gender difference should be taken into consideration when evaluating fibre CSA in myositis.

A high incidence of disease flares in an open pilot study of infliximab in patients with refractory inflammatory myopathies.

OBJECTIVE: To investigate the effect of the tumour necrosis factor (TNF) blocking agent infliximab in patients with treatment-resistant inflammatory myopathies. METHODS: A total of 13 patients with refractory polymyositis (PM), dermatomyositis (DM), or inclusion body myositis (IBM) were treated with 4 infliximab infusions (5 mg/kg body weight) over 14 weeks. Outcome measures included myositis disease activity score with improvement defined according to The International Myositis Assessment and Clinical Studies Group (IMACS), and MRI. Repeated muscles biopsies were investigated for cellular infiltrates, major histocompatibility complex (MHC) class I and II, TNF, interleukin (IL)1alpha, IL6, high mobility group box chromosomal protein 1 (HMGB-1), interferon gamma (IFNgamma), myxovirus resistance protein A (MxA) and membrane attack complex (MAC) expression. Type I IFN activity was analysed in sera. RESULTS: Nine patients completed the study. Three patients discontinued due to adverse events and one due to a discovered malignancy. Three of the completers improved by >or=20% in three or more variables of the disease activity core set, four were unchanged and two worsened >or=30%. No patient improved in muscle strength by manual muscle test. At baseline, two completers had signs of muscle inflammation by MRI, and five at follow-up. T lymphocytes, macrophages, cytokine expression and MAC deposition in muscle biopsies were still evident after treatment. Type I IFN activity was increased after treatment. CONCLUSIONS: Infliximab treatment was not effective in refractory inflammatory myopathies. In view of radiological and clinical worsening, and activation of the type I IFN system in several cases, infliximab is not an alternative treatment in patients with treatment-resistant myositis.

Interstitial lung disease, a common manifestation of newly diagnosed polymyositis and dermatomyositis.

OBJECTIVES: To estimate the prevalence and predictors of interstitial lung disease in newly diagnosed polymyositis and dermatomyositis. METHODS: A prospective study in which consecutive patients with newly diagnosed poly- and dermatomyositis, regardless of clinical symptoms of pulmonary disease, were investigated with chest x ray, high resolution computed tomography (HRCT), pulmonary function tests, and biochemical and autoantibody analysis. Patients with inclusion body myositis, malignancy, other defined inflammatory connective tissue diseases (CTDs), or antibody profile indicating other CTDs were excluded. RESULTS: Between March 1998 and September 2000, 26 new cases of poly- or dermatomyositis were diagnosed; 17 of those patients were included in the study. Interstitial lung disease (ILD), defined as radiological signs on chest x ray examination/HRCT or restrictive ventilatory defect, were found in 11 (65%) patients and were more common in men than in women. Arthritis and occurrence of anti-Jo-1 antibodies were found more often in patients with ILD than in those without. There was no statistically significant association between respiratory symptoms, other serological or laboratory variables and ILD. CONCLUSIONS: ILD is a common early manifestation in patients with poly- and dermatomyositis and is not always related to clinical symptoms. Chest x ray examination, HRCT, pulmonary function tests, and analysis of anti-Jo-1 antibodies should be included in the initial investigation of patients with myositis regardless of respiratory symptoms.