Genotype-guided diagnostic reassessment after exome sequencing in neuromuscular disorders: experiences with a two-step approach.

BACKGROUND AND PURPOSE: Next-generation sequencing has greatly improved the diagnostic success rates for genetic neuromuscular disorders (NMDs). Nevertheless, most patients still remain undiagnosed, and there is a need to maximize the diagnostic yield. METHODS: A retrospective study was conducted on 72 patients with NMDs who underwent exome sequencing (ES), partly followed by genotype-guided diagnostic reassessment and secondary investigations. The diagnostic yields that would have been achieved by appropriately chosen narrow and comprehensive gene panels were also analysed. RESULTS: The initial diagnostic yield of ES was 30.6% (n = 22/72 patients). In an additional 15.3% of patients (n = 11/72) ES results were of unknown clinical significance. After genotype-guided diagnostic reassessment and complementary investigations, the yield was increased to 37.5% (n = 27/72). Compared to ES, targeted gene panels (<25 kilobases) reached a diagnostic yield of 22.2% (n = 16/72), whereas comprehensive gene panels achieved 34.7% (n = 25/72). CONCLUSION: Exome sequencing allows the detection of pathogenic variants missed by (narrowly) targeted gene panel approaches. Diagnostic reassessment after genetic testing further enhances the diagnostic outcomes for NMDs.

Hereditary spastic paraplegia caused by compound heterozygous mutations outside the motor domain of the KIF1A gene.

BACKGROUND AND PURPOSE: Hereditary spastic paraplegia is a clinically and genetically heterogeneous group of rare, inherited disorders causing an upper motor neuron syndrome with (complex) or without (pure) additional neurological symptoms. Mutations in the KIF1A gene have already been associated with recessive and dominant forms of hereditary spastic paraplegia (SPG30) in a few cases. METHODS: All family members included in the study were examined neurologically. Whole-exome sequencing was used in affected individuals to identify the responsible candidate gene. Conventional Sanger sequencing was conducted to validate familial segregation. RESULTS: A family of Macedonian origin with two affected siblings, one with slowly progressive and the other one with a more complex and rapidly progressing hereditary spastic paraplegia is reported. In both affected individuals, two novel pathogenic mutations outside the motor domain of the KIF1A gene were found (NM_001244008.1:c.2909G>A, p.Arg970His and c.1214dup, p.Asn405Lysfs*40) that segregate with the disease within the family establishing the diagnosis of autosomal recessive SPG30. CONCLUSIONS: This report provides the first evidence that mutations outside the motor domain of the gene can cause (recessive) SPG30 and extends the genotype-phenotype association for KIF1A-related diseases.

[Adrenoleukodystrophy mimicking multiple sclerosis]. / Adrenoleukodystrophie kann Multiple Sklerose imitieren.

The article describes the development of symptoms in a 40-year-old female patient who is a symptomatic carrier of X-linked adrenoleucodystrophy (ALD). ALD is characterized by impaired peroxisomal beta-oxidation of very long chain fatty acids and is associated with mutations of the ALD gene, resulting in a defective peroxisomal membrane-transport protein. Our patient's symptoms are identical to those found in multiple sclerosis, showing spastic paraparesis of the lower limbs with marked sensory deficits, visual disturbances in the right eye, and bladder difficulties. Visual and auditory evoked potentials were pathological, and a cranial MRI revealed multiple periventrical white-matter lesions. We found increased intrathecal immunoglobulin production. Diagnosis was established by high concentrations of very long chain fatty acids in serum and in dermal fibroblasts after the same was found in our patient's son. In familial multiple sclerosis, ALD should be excluded in male and female patients.

Giant-cell tumor of the synovial membrane: localized nodular synovitis in the knee joint.

Giant-cell tumor of the synovia is a benign neoplasm characterized histologically by proliferating histiocytes bearing lipids and hemosiderin intermingled with a variable number of multinuclear giant cells. Areas of predilection are the hand, and in the case of synovial joints, the knee joint is particularly affected. Clinically, patients have signs of mechanical derangement and, with the knee joint, meniscal symptoms and locking are often present. Joint effusion without previous trauma is another typical finding. Diagnosis is carried out by radiographic tools and has to be confirmed histologically. Giant-cell tumor of the synovia is treated by local excision either by arthroscopy or by arthrotomy. To our knowledge, the case we present is the largest giant-cell tumor of the synovia in the knee joint ever described in literature.

Involution and neoplasms of the thymus.

Mammalian ontogenesis and postnatal histogenesis involves the dynamic and appropriate interaction of two growth related phenomena: progression and regression. The thymus gland is the organ of the mammalian body that exhibits the most profound involution during normal postnatal histogenesis. Involution of the thymus can be compared to similarly regressive processes during the ontogeny of holometabolic insects, as well as to the spontaneous regression of neoplasms. It can be expected that in the future a better understanding of neoplastic regression will result from the comparison of ontogenetic processes from taxonomically far-removed regressive processes, and the evaluation of various factors that promote progression and regression. Furthermore the thymus, as the key organ of the cellular arm of the mammalian immune system, is intimately involved in the determination of the eventual outcome of malignant neoplastic disease. Primary anterior mediastinal tumors comprise a diverse group of neoplasms accounting for 50% of all mediastinal tumor lesions. Thymic carcinomas and thymomas are epithelial neoplasms which can be divided according to clinical behavior into various biological subtypes. A review of the nature of thymic neoplasms, a description of physiologic and accidental involution of the thymus gland, and a discussion of various paraneoplastic disorders, including myesthenia gravis, associated with thymic neoplasms are the aims of this article.

Neoplasms during the progression of pregnancy.

Restricting this review to dealing with pregnancy and its interaction with neoplasms limits us to the child-bearing years. Neoplasms may appear at all stages of species with true tissues and the incidence of malignancy in pregnancy is estimated to be 1:1,000. Almost 50% of these tumors are cervical cancers, followed by breast cancer, with an incidence of approximately 0.03%. The pregnant woman, in the same person, exhibits controlled growth (the pregnancy) and uncontrolled growth (the malignancy). In younger women, the neoplasms represent early stages of biological development and seem to arise practically from all maternal tissues. Geriatric changes in the neoplastic growth processes are missing. This article encompasses a review of the integration of neoplasms, the maternal body, the fetus and the placenta. The morphological and biochemical integration of the different processes is diversified. Mainly, we would like to address the interaction between pregnancy and common human malignancies like breast, cervix, and melanoma, but we will also review rare neoplastic complications. This way it is possible to treat the combined growth processes as they evolve from the initiated sperm, the ovum and continue via the placental development. These processes lead to the fetus and, in the pathological sense, to childhood complications, though most cases develop only portions of the process.

Growth inhibitors in the treatment of malignant neoplasms.

Growth inhibitors are an integral part of the regulatory mechanisms involved in the growth and differentiation of cells and tissues. Aberration in the response to growth inhibitors leads to the escape of the cell from the cell cycle control mechanisms and may lead to the development of malignancies. The inactivation of tumor suppressor genes, activation of oncogenes leads to the acquisition of an invasive and increasingly malignant immunophenotype and secretory profile by transformed cells. The commencement of the complex process of carcinogenesis, and subsequent, rapid tumor growth and progression of mammalian neoplasms depends upon the continuous de novo formation of capillaries (angiogenesis). The generation of a malignant, invasive cellular immunophenotype (CIP) and distant metastases, as aspects of tumor progression, are also NRA-dependent processes. Specific molecules with cytostatic/cytotoxic growth inhibitory effects represent a very diverse group of factors. Growth inhibitors may regulate the cell cycle at various levels, and growth inhibitors comprise a heterogeneous group of agents including cytokines, growth factors, steroid hormones, etc. The phenomenon of multidrug resistance to a wide spectrum of cytostatic/cytotoxic agents has posed a major difficulty in the effective chemotherapeutical treatment of cancer patients. The development of novel therapeutic regimens should be based on the observations of the growth inhibitory profile of the particular malignancy, in addition to its immunophenotype and genotype, and the devisement of 'individualized' combinations of factors, including gene and immuno-therapeutical options, targeting different aspects of the malignant disease.

Interdonor variability of platelet response to thrombin receptor activation: influence of PlA2 polymorphism.

Considering that platelet response to thrombin receptor activation might be critical for the development of arterial thrombosis, we measured the dense granule release under stimulation by the thrombin receptor activating peptide (TRAP) in a series of 102 healthy volunteers. The threshold TRAP concentration which initiated a secretion ranged from 3 to 20 microM. A good concordance (79%, k=0.677) between two tests performed at a 1 month interval indicated that platelet response to thrombin receptor activation was characteristic of each individual donor. Since the threshold concentration required to initiate secretion corresponded to the threshold concentration which induced a biphasic aggregation, all volunteers were genotyped for the PlA2 polymorphism, the Pro33 variant of GPIIIa. Platelets from subjects with the PlA2 polymorphism required higher TRAP concentrations to aggregate than those from subjects with no PlA2 allele (P=0.0012). However, they also required a higher ADP concentration to aggregate. In order to exclude any influence of GPIIIa polymorphism on TRAP-induced secretion, we studied the variability of platelet response to TRAP among the 77 individuals with no PlA2 allele, and found the same interdonor variability with the same distribution of threshold TRAP concentrations as for the 102 individuals. The results suggest that (i) platelet secretion in response to thrombin receptor activation could be a genetically controlled phenotype independent of the GPIIIa polymorphism; (ii) the PlA2 polymorphism is associated with platelet hypoaggregability.

Improvements in solubility and stability of thalidomide upon complexation with hydroxypropyl-beta-cyclodextrin.

Thalidomide is in clinical use for the treatment of graft-versus-host disease in leukemia patients after bone marrow transplant. Low levels of the drug in plasma after oral administration have made an intravenous thalidomide formulation desirable. Thalidomide, however, is sparingly soluble in aqueous solution (50 micrograms/mL) and unstable. Complexation with hydroxypropyl-beta-cyclodextrin has significantly improved the aqueous solubility and stability of thalidomide. Results obtained with HPLC and 1H NMR spectrometry have demonstrated that the solubility is increased to 1.7 mg/mL and the half-life of a diluted solution is extended from 2.1 to 4.1 h. Hence, an intravenous thalidomide-hydroxypropyl- beta-cyclodextrin solution has the potential to significantly improve current therapy for graft-versus-host disease by providing sustained high levels of drug in the plasma.

Perception of figure orientation by nonambulatory children with profound mental retardation.

An habituation-dishabituation procedure was used to study perception of changes in figure orientation by nonambulatory children with profound mental retardation. On each of 3 days, 16 participants were given familiarization trials with either a vertical or an horizontal pattern, followed by test trials with the familiarized stimulus and one of three novel stimuli--a 90 degree or 45 degree rotation of the pattern, or a rearrangement of the pattern elements into a square. Visual fixation times decreased over familiarization trials. Fixation times on test trials were longer for novel than for familiar stimuli, indicating that these children discriminated changes in form and orientation.

The benefit of lithium carbonate adjunct in refractory depression--fact or fiction?

Our group attempted to validate previous claims of rapid success with lithium carbonate adjunct therapy in antidepressant-resistant depression. Seven depressed patients volunteered for a study of placebo controlled, double-blind design. During their treatment on general hospital psychiatric wards, these patients received antidepressant medication for a period of at least 21 days. While continuing to receive antidepressant medication after the 21 day period, four of the patients then received lithium carbonate, and three received placebo. This combination pharmacological therapy continued for a 48 hour period. After this time, six of the seven patients showed no significant improvement. The remaining patient, who had received lithium carbonate, improved markedly over the 48 hour period. However, this patient relapsed within one week. A review of the two most extensive studies claiming significant results with the lithium carbonate adjunct therapy was performed. We feel that they, as presented, leave serious doubt as to the validity of their conclusions. We conclude that on the basis of our work up to this point in time and the analysis of previous reports claiming otherwise, no valid evidence exists for a consistent therapeutic effect of lithium carbonate adjunct in antidepressant-resistant depression. It was also found that methodologic contamination necessitated the exclusion of an additional six patients from the double-blind trial. We conclude that in order to objectively examine the rapidly expanding field of biological psychiatry, teaching of clinical staff in basic research procedure should be stressed as a part of routine ward orientation.

Uses of the Bayley mental scale with nonambulatory profoundly mentally retarded children.

A set of 45 mental scale items from the Bayley Scales of Infant Development was administered by two testers to 33 nonambulatory, profoundly mentally retarded subjects. Interobserver agreement was high for overall scores and for 42 of 45 individual items. Total number of items passed was significantly correlated with activity level. Division of these items into a stimulus set and a response set provided a qualitative description of performance that supported previous research indicating that these individuals have deficits in exploratory motivation and perceptual curiosity. Bayley raw scores predicted success in conditioning programs, suggesting that this test is useful for educational programming.