Ocrelizumab quantitation by liquid chromatography-tandem mass spectrometry.

Introduction: Ocrelizumab is a monoclonal anti-CD20 antibody approved for the treatment of multiple sclerosis (MS). The clinical value of therapeutic drug monitoring (TDM) for this antibody in treatment of MS is unknown, and an adequately specific and precise quantitation method for ocrelizumab in patient serum could facilitate investigation. Liquid chromatography-tandem mass spectrometry (LC-MS/MS)-based quantitation methods have been shown to have higher analytic specificity and precision than enzyme-linked immunosorbent assays. Objectives: To establish and validate an LC-MS/MS-based quantitation method for ocrelizumab. Methods: We present an LC-MS/MS-based quantitation method using immunocapture purification followed by trypsinization and analysis by a triple quadrupole mass analyzer obtaining results within the same day. Results: We found that the ocrelizumab peptide GLEWVGAIYPGNGDTSYNQK (Q1/Q3 Quantifier ion: 723.683+/590.77 y112+ Qualifier ion: 723.683+/672.30 y122+) can be used for quantitation and thereby developed a method for quantifying ocrelizumab in human serum with a quantitation range of 1.56 to 200 µg/mL. The method was validated in accordance with EMA requirements in terms of selectivity, carry-over, lower limit of quantitation, calibration curve, accuracy, precision and matrix effect. Ocrelizumab serum concentrations were measured in three MS patients treated with ocrelizumab, immediately before and after ocrelizumab infusion, with additional sampling after 2, 4, 8 and 12 weeks. Measured serum concentrations of ocrelizumab showed expected values for both Cmax and drug half-life over the sampled time period. Conclusion: We have established a reliable quantitation method for serum ocrelizumab that can be applied in clinical studies, facilitating the evaluation of ocrelizumab TDM in MS.

Identification of an iron-responsive subtype in two children diagnosed with relapsing-remitting multiple sclerosis using whole exome sequencing.

BACKGROUND: Multiple sclerosis is a disorder related to demyelination of axons. Iron is an essential cofactor in myelin synthesis. Previously, we described two children (males of mixed ancestry) with relapsing-remitting multiple sclerosis (RRMS) where long-term remission was achieved by regular iron supplementation. A genetic defect in iron metabolism was postulated, suggesting that more advanced genetic studies could shed new light on disease pathophysiology related to iron. METHODS: Whole exome sequencing (WES) was performed to identify causal pathways. Blood tests were performed over a 10 year period to monitor the long-term effect of a supplementation regimen. Clinical wellbeing was assessed quarterly by a pediatric neurologist and regular feedback was obtained from the schoolteachers. RESULTS: WES revealed gene variants involved in iron absorption and transport, in the transmembrane protease, serine 6 (TMPRSS6) and transferrin (TF) genes; multiple genetic variants in CUBN, which encodes cubilin (a receptor involved in the absorption of vitamin B12 as well as the reabsorption of transferrin-bound iron and vitamin D in the kidneys); SLC25A37 (involved in iron transport into mitochondria) and CD163 (a scavenger receptor involved in hemorrhage resolution). Variants were also found in COQ3, involved with synthesis of Coenzyme Q10 in mitochondria. Neither of the children had the HLA-DRB1*1501 allele associated with increased genetic risk for MS, suggesting that the genetic contribution of iron-related genetic variants may be instrumental in childhood MS. In both children the RRMS has remained stable without activity over the last 10 years since initiation of nutritional supplementation and maintenance of normal iron levels, confirming the role of iron deficiency in disease pathogenesis in these patients. CONCLUSION: Our findings highlight the potential value of WES to identify heritable risk factors that could affect the reabsorption of transferrin-bound iron in the kidneys causing sustained iron loss, together with inhibition of vitamin B12 absorption and vitamin D reabsorption (CUBN) and iron transport into mitochondria (SLC25A37) as the sole site of heme synthesis. This supports a model for RRMS in children with an apparent iron-deficient biochemical subtype of MS, with oligodendrocyte cell death and impaired myelination possibly caused by deficits of energy- and antioxidant capacity in mitochondria.

Landmark studies in neuro-ophthalmology

@#High-quality clinical evidence, derived from well-designed and implemented clinical trials, serves to advance clinical care and to allow physicians to provide the most effective treatments to their patients. The field of ophthalmology, including the subspecialty of neuro-ophthalmology, abounds with such high-quality clinical trials that provide Level 1 clinical evidence. This review article summarizes the research design, key findings, and clinical relevance of select monumental clinical studies in neuro-ophthalmology with the primary goal of providing the readers with the rationale for current standard of care of various neuro-ophthalmic diseases. This includes the Optic Neuritis Treatment Trial, Ischemic Optic Neuropathy Decompression Trial, Idiopathic Intracranial Hypertension Treatment Trial, Rescue of Hereditary Optic Disease Outpatient Study, and Controlled High-Risk Avonex® Multiple Sclerosis Study

A long-term follow-up study using IPMSSG criteria in children with CNS demyelination.

OBJECTIVE: To evaluate the practical application of International Pediatrics Multiple Sclerosis study group definitions in children with inflammatory demyelination of the central nervous system and to identify predictors of multiple sclerosis. METHODS: Baseline data on 123 children with a first episode of acute central nervous system demyelination were collected. The initial diagnosis according to the International Pediatrics Multiple Sclerosis study group was recorded and compared with final diagnosis. RESULTS: Forty-seven (38.2%) children met International Pediatrics Multiple Sclerosis study group criteria for acute disseminated encephalomyelitis and 67 (54.4%) had clinically isolated syndrome at the initial presentation. Four (3.2%) had the diagnosis of neuromyelitis optica and five (4%) did not meet any specific diagnosis per the study group criteria. Clinical follow-up was available on 118 of 123 children (95.9%), with a median of 61.5 months (quartile range 23, 110 months). Conversion from clinically isolated syndrome to multiple sclerosis occurred in 26 of 67 children (38.8%); acute disseminated encephalomyelitis to multiple sclerosis occurred in 4 of 47 children (8.5%). Adjusted multivariate logistic regression analysis for an outcome of future development of multiple sclerosis showed the following predictors: female gender (odds ratio 12.44; 95% confidence interval 1.03-149.3); initial diagnosis of monofocal brain stem or hemispheric dysfunction (odds ratio 24.57; 95% confidence interval 3.06-196.78); and Callen magnetic resonance imaging criteria if met (odds ratio 122.45; 95% confidence interval 16.57-904.57). CONCLUSION: International Pediatrics Multiple Sclerosis study group criteria affirm that children with initial clinically isolated syndrome are more likely to develop future multiple sclerosis compared with those with an acute disseminated encephalomyelitis initial diagnosis. In addition, female gender, brain stem or hemispheric involvement, and Callen magnetic resonance imaging criteria predict the diagnosis of multiple sclerosis.