Extended interval dosing of ocrelizumab modifies the repopulation of B cells without altering the clinical efficacy in multiple sclerosis.

BACKGROUND: Recent studies suggest that extended interval dosing of ocrelizumab, an anti-B cell therapy, does not affect its clinical effectiveness in most patients with multiple sclerosis (MS). However, it remains to be established whether certain B cell subsets are differentially repopulated after different dosing intervals and whether these subsets relate to clinical efficacy. METHODS: We performed high-dimensional single-cell characterization of the peripheral immune landscape of patients with MS after standard (SID; n = 43) or extended interval dosing (EID; n = 37) of ocrelizumab and in non-ocrelizumab-treated (control group, CG; n = 28) patients with MS, using mass cytometry by time of flight (CyTOF). RESULTS: The first B cells that repopulate after both ocrelizumab dosing schemes were immature, transitional and regulatory CD1d+ CD5+ B cells. In addition, we observed a higher percentage of transitional, naïve and regulatory B cells after EID in comparison with SID, but not of memory B cells or plasmablasts. The majority of repopulated B cell subsets showed an increased migratory phenotype, characterized by higher expression of CD49d, CD11a, CD54 and CD162. Interestingly, after EID, repopulated B cells expressed increased CD20 levels compared to B cells in CG and after SID, which was associated with a delayed repopulation of B cells after a subsequent ocrelizumab infusion. Finally, the number of/changes in B cell subsets after both dosing schemes did not correlate with any relapses nor progression of the disease. CONCLUSIONS: Taken together, our data highlight that extending the dosing interval of ocrelizumab does not lead to increased repopulation of effector B cells. We show that the increase of CD20 expression on B cell subsets in EID might lead to longer depletion or less repopulation of B cells after the next infusion of ocrelizumab. Lastly, even though extending the ocrelizumab interval dosing alters B cell repopulation, it does not affect the clinical efficacy of ocrelizumab in our cohort of patients with MS.

Pre-analytical stability of serum biomarkers for neurological disease: neurofilament-light, glial fibrillary acidic protein and contactin-1.

OBJECTIVES: Neurofilament-light (NfL), glial fibrillary acidic protein (GFAP) and contactin-1 (CNTN1) are blood-based biomarkers that could contribute to monitoring and prediction of disease and treatment outcomes in neurological diseases. Pre-analytical sample handling might affect results, which could be disease-dependent. We tested common handling variations in serum of volunteers as well as in a defined group of patients with multiple sclerosis (pwMS). METHODS: Sample sets from 5 pwMS and 5 volunteers at the outpatient clinic were collected per experiment. We investigated the effect of the following variables: collection tube type, delayed centrifugation, centrifugation temperature, delayed storage after centrifugation and freeze-thawing. NfL and GFAP were measured by Simoa and CNTN1 by Luminex. A median recovery of 90-110% was considered stable. RESULTS: For most pre-analytical variables, serum NfL and CNTN1 levels remained unaffected. In the total group, NfL levels increased (121%) after 6 h of delay at 2-8 °C until centrifugation, while no significant changes were observed after 24 h delay at room temperature (RT). In pwMS specifically, CNTN1 levels increased from additional freeze-thaw cycles number 2 to 4 (111%-141%), whereas volunteer levels remained stable. GFAP showed good stability for all pre-analytical variables. CONCLUSIONS: Overall, the serum biomarkers tested were relatively unaffected by variations in sample handling. For serum NfL, we recommend storage at RT before centrifugation at 2-8 °C up to 6 h or at RT up to 24 h. For serum CNTN1, we advise a maximum of two freeze-thaw cycles. Our results confirm and expand on recently launched consensus standardized operating procedures.

Project Y: The search for clues explaining phenotype variability in MS.

BACKGROUND: To study phenotypic variability in MS patients, well-defined unbiased cohort studies are necessary. The most common and probably most important confounding factor when studying disease phenotype in MS is age. OBJECTIVE: To describe study design and subject characteristics of a unique birth cohort (Project Y). The overall aim of Project Y is to identify determinants associated with phenotypic variability in MS, eliminating the possibility of confounding by age. METHODS: Project Y is a population-based cross-sectional study of all people with MS born in the Netherlands in 1966. Patients and healthy controls were subjected to comprehensive examinations: functional and static imaging, physical and cognitive measurements, and lifestyle factors early and later in life. In addition body fluids were collected and stored for future biomarker research. RESULTS: 452 eligible MS patients were identified. Between December 2017 and January 2021, 367 MS patients and 125 healthy controls participated. The total number of identified cases results in a current prevalence of at least 189/100.000 for people born in the year 1966 in The Netherlands. CONCLUSION: Project Y is a unique cohort designed to identify factors associated with phenotypic variability in MS patients without the confounding effects of age. This first description of the Project Y cohort indicates that the prevalence of MS in the Netherlands might be higher than previously presumed. Various studies using Project Y data are ongoing and results will be published in upcoming years.

Serum Neurofilament Light Association With Progression in Natalizumab-Treated Patients With Relapsing-Remitting Multiple Sclerosis.

BACKGROUND AND OBJECTIVES: To investigate the potential of serum neurofilament light (NfL) to reflect or predict progression mostly independent of acute inflammatory disease activity in patients with relapsing-remitting multiple sclerosis (RRMS) treated with natalizumab. METHODS: Patients were selected from a prospective observational cohort study initiated in 2006 at the VU University Medical Center Amsterdam, the Netherlands, including patients with RRMS treated with natalizumab. Selection criteria included an age of 18 years or older and a minimum follow-up of 3 years from natalizumab initiation. Clinical and MRI assessments were performed on a yearly basis, and serum NfL was measured at 5 time points during the follow-up, including on the day of natalizumab initiation (baseline), 3 months, 1 year, and 2 years after natalizumab initiation, and on last follow-up visit. Using general linear regression models, we compared the longitudinal dynamics of NfL between patients with and without confirmed Expanded Disability Status Scale (EDSS) progression between year 1 visit and last follow-up, and between individuals with and without EDSS+ progression, a composite endpoint including the EDSS, 9-hole peg test, and timed 25-foot walk. RESULTS: Eighty-nine natalizumab-treated patients with RRMS were included. Median follow-up time was 5.2 years (interquartile range [IQR] 4.3-6.7, range 3.0-11.0) after natalizumab initiation, mean age at time of natalizumab initiation was 36.9 years (SD 8.5), and median disease duration was 7.4 years (IQR 3.8-12.1). Between year 1 and the last follow-up, 28/89 (31.5%) individuals showed confirmed EDSS progression. Data for the EDSS+ endpoint was available for 73 out of the 89 patients and 35/73 (47.9%) showed confirmed EDSS+ progression. We observed a significant reduction in NfL levels 3 months after natalizumab initiation, which reached its nadir of close to 50% of baseline levels 1 year after treatment initiation. We found no difference in the longitudinal dynamics of NfL in progressors vs nonprogressors. NfL levels at baseline and 1 year after natalizumab initiation did not predict progression at last follow-up. CONCLUSION: In our cohort of natalizumab-treated patients with RRMS, NfL fails to capture or predict progression that occurs largely independently of clinical or radiologic signs of acute focal inflammatory disease activity. Additional biomarkers may thus be needed to monitor progression in these patients. CLASSIFICATION OF EVIDENCE: This study provides Class II evidence that serum NfL levels are not associated with disease progression in natalizumab-treated patients with RRMS.

Mild progressive multifocal leukoencephalopathy after switching from natalizumab to ocrelizumab.

OBJECTIVE: To describe the disease course of carryover progressive multifocal leukoencephalopathy (PML) after switching from natalizumab to ocrelizumab in 2 patients with relapsing-remitting MS. METHODS: Two case reports with 1 year of follow-up and retrospective longitudinal measurements of serum neurofilament light (NfL) levels and B-cells. RESULTS: PML was diagnosed 78 days (case 1) and 97 days (case 2) after discontinuation of natalizumab. Both patients developed mild immune reconstitution inflammatory syndrome (IRIS) despite B-cell depletion caused by ocrelizumab. NfL levels increased in both patients during PML-IRIS. PML-IRIS lesions stabilized after treatment with mefloquine and mirtazapine, followed by methylprednisolone, and both patients continued therapy with ocrelizumab when B-cells started to repopulate. CONCLUSIONS: The clinical course of carryover PML was mild in both patients, suggesting that B-cell depletion possibly did not aggravate PML-IRIS in these 2 patients.

Thyroid Gland 18F-FDG Uptake in Neurofibromatosis Type 1.

PURPOSE: To investigate thyroid gland characteristics on 18F-FDG positron emission tomography/computed tomography (PET/CT) imaging in patients with neurofibromatosis type 1 (NF1). SUBJECTS AND METHODS: Thyroid gland characteristics of patients with a clinical diagnosis of NF1 who underwent 18F-FDG PET/CT imaging for the first time to distinguish benign neurofibroma from malignant peripheral nerve sheath tumor (MPNST) at our institution (n = 69) were compared to PET/CT imaging of sarcoidosis (n = 25) and early stage lung cancer (T1N0M0 tumors, n = 15) patients. RESULTS: Two NF1 patients (3%) showed a diffuse 18F-FDG uptake in the thyroid gland, 2 patients (3%) had an irregular uptake, and 7 patients (10%) had a focal uptake. Among the sarcoidosis patients, 1 showed a diffuse uptake (4%) and 1 had an irregular uptake (4%). In the early stage lung cancer group, 1 patient showed a diffuse uptake (7%) and 1 had a focal uptake (7%). NF1 patients had larger mean thyroid volume and mean SUVmax compared to sarcoidosis patients but not compared to early stage lung cancer patients. Four NF1 patients were diagnosed with multinodular goiter, 2 patients were diagnosed with benign chronic lymphocytic thyroiditis, 1 patient had metastasis to the thyroid, and 1 patient had medullary thyroid cancer. CONCLUSION: Even though NF1 patients did not show an increased risk of thyroid incidentaloma on PET/CT compared to previous studies on non-thyroid cancer patients, the incidence shows that awareness of possible thyroid disease is important.