Differential effects of anti-CD20 therapy on CD4 and CD8 T cells and implication of CD20-expressing CD8 T cells in MS disease activity.

A small proportion of multiple sclerosis (MS) patients develop new disease activity soon after starting anti-CD20 therapy. This activity does not recur with further dosing, possibly reflecting deeper depletion of CD20-expressing cells with repeat infusions. We assessed cellular immune profiles and their association with transient disease activity following anti-CD20 initiation as a window into relapsing disease biology. Peripheral blood mononuclear cells from independent discovery and validation cohorts of MS patients initiating ocrelizumab were assessed for phenotypic and functional profiles using multiparametric flow cytometry. Pretreatment CD20-expressing T cells, especially CD20dimCD8+ T cells with a highly inflammatory and central nervous system (CNS)-homing phenotype, were significantly inversely correlated with pretreatment MRI gadolinium-lesion counts, and also predictive of early disease activity observed after anti-CD20 initiation. Direct removal of pretreatment proinflammatory CD20dimCD8+ T cells had a greater contribution to treatment-associated changes in the CD8+ T cell pool than was the case for CD4+ T cells. Early disease activity following anti-CD20 initiation was not associated with reconstituting CD20dimCD8+ T cells, which were less proinflammatory compared with pretreatment. Similarly, this disease activity did not correlate with early reconstituting B cells, which were predominantly transitional CD19+CD24highCD38high with a more anti-inflammatory profile. We provide insights into the mode-of-action of anti-CD20 and highlight a potential role for CD20dimCD8+ T cells in MS relapse biology; their strong inverse correlation with both pretreatment and early posttreatment disease activity suggests that CD20-expressing CD8+ T cells leaving the circulation (possibly to the CNS) play a particularly early role in the immune cascades involved in relapse development.

Resolving variable maternal D typing using serology and genotyping in selected prenatal patients.

BACKGROUND: RhIG prophylaxis for D- pregnant women prevents hemolytic disease of the newborn and typically depends on results of serologic D typing. Interpretation and follow-up of weak D serology is variable. Recent recommendations promote genotyping for RHD status determination in those with weak D serology. Canadian Blood Services performs comprehensive serologic prenatal testing in four provinces. Genotyping is used to determine D typing in patients with weak D. STUDY DESIGN AND METHODS: A serologic algorithm identified which patients require genotyping for RHD determination. Genotyping was performed on one of two commercially available platforms. RESULTS: Only 0.4% of D- patients met criteria for genotyping. Sixty-one percent were weak D Type 1, 2, or 3. Thirty percent had a partial or weak D other than Type 1, 2, or 3. Eleven had variants which remained unresolved. Seventeen were D+ and four were D-. CONCLUSIONS: Genotyping of patients with weak D serology led to an identified genotype in most patients. RhIG administration was avoided in 66% who were weak D Type 1, 2, or 3 or were D+. The use of a serologic algorithm to select patients for RHD genotyping identifies a majority of patients with weak D types not at risk for alloimmunization. This approach limits the number of genotyping investigations and the cost of providing classification for weak D types.

The prevalence of anti-K in Canadian prenatal patients.

BACKGROUND: Anti-KEL1(K) is a major cause of hemolytic disease of the fetus and newborn. We utilized data from prenatal testing of patients in Western Canada to determine the frequency of anti-K. In Manitoba, we evaluated the frequency of transfusion as the likely cause for alloimmunization. We reviewed international practices to prevent alloimmunization. STUDY DESIGN AND METHODS: Prenatal patients undergo antibody screening using an automated testing platform and uniform testing algorithm. Data on the frequency of antibodies, transfusion history, and donor K typing were extracted from the relevant databases at Canadian Blood Services. National standards were reviewed with the help of local experts. RESULTS: Anti-K was found in 397 of 390,193 patients from 2011 to 2013 (1.02 per 1000) and was the second most frequent antibody after anti-E. In Manitoba, 26 of 75 (35%) anti-K patients had received transfusions in the province since 2001; 14 of the 26 (54%) had received at least one K+ RBC unit and three had received all K- units, while in nine, donor K typing was incomplete. Only eight of the 26 had previous pregnancies, three with K+ partners. International practice varies; however, prophylactic use of matched or K- units is standard in many European countries. CONCLUSIONS: Anti-K was found in 0.1% of prenatal patients. Although our data on the history of transfusion are incomplete, they demonstrate that transfusion with a K+ unit is a major cause of alloimmunization. Given advances in phenotyping and genotyping technologies, prophylactic matching should be considered in Canada.

Emerging Cerebrospinal Fluid Biomarkers of Disease Activity and Progression in Multiple Sclerosis.

Importance: Biomarkers distinguishing nonrelapsing progressive disease biology from relapsing biology in multiple sclerosis (MS) are lacking. Cerebrospinal fluid (CSF) is an accessible fluid that most closely reflects central nervous system biology. Objective: To identify CSF biological measures associated with progressive MS pathobiology. Design, Setting, and Participants: This cohort study assessed data from 2 prospective MS cohorts: a test cohort provided serial CSF, clinical, and imaging assessments in a multicenter study of patients with relapsing MS (RMS) or primary progressive MS (PPMS) who were initiating anti-CD20 treatment (recruitment: 2016-2018; analysis: 2020-2023). A single-site confirmation cohort was used to assess CSF at baseline and long-term (>10 year) clinical follow-up (analysis: 2022-2023). Exposures: Test-cohort participants initiated standard-of-care ocrelizumab treatment. Confirmation-cohort participants were untreated or received standard-of-care disease-modifying MS therapies. Main Outcomes and Measures: Twenty-five CSF markers, including neurofilament light chain, neurofilament heavy chain, and glial fibrillary acid protein (GFAP); 24-week confirmed disability progression (CDP24); and brain magnetic resonance imaging measures reflecting focal injury, tissue loss, and progressive biology (slowly expanding lesions [SELs]). Results: The test cohort (n = 131) included 100 patients with RMS (mean [SD] age, 36.6 [10.4] years; 68 [68%] female and 32 [32%] male; Expanded Disability Status Scale [EDSS] score, 0-5.5), and 31 patients with PPMS (mean [SD] age, 44.9 [7.4] years; 15 [48%] female and 16 [52%] male; EDSS score, 3.0-6.5). The confirmation cohort (n = 68) included 41 patients with RMS and 27 with PPMS enrolled at diagnosis (age, 40 years [range, 20-61 years]; 47 [69%] female and 21 [31%] male). In the test cohort, GFAP was correlated with SEL count (r = 0.33), greater proportion of T2 lesion volume from SELs (r = 0.24), and lower T1-weighted intensity within SELs (r = -0.33) but not with acute inflammatory measures. Neurofilament heavy chain was correlated with SEL count (r = 0.25) and lower T1-weighted intensity within SELs (r = -0.28). Immune markers correlated with measures of acute inflammation and, unlike GFAP, were impacted by anti-CD20. In the confirmation cohort, higher baseline CSF GFAP levels were associated with long-term CDP24 (hazard ratio, 2.1; 95% CI, 1.3-3.4; P = .002). Conclusions and Relevance: In this study, activated glial markers (in particular GFAP) and neurofilament heavy chain were associated specifically with nonrelapsing progressive disease outcomes (independent of acute inflammatory activity). Elevated CSF GFAP was associated with long-term MS disease progression.

Changes in Fatigue and Cognition in Patients with Relapsing Forms of Multiple Sclerosis Treated with Natalizumab: The ENER-G Study.

Fatigue and cognitive impairment are debilitating features of multiple sclerosis (MS). ENER-G was a 12-month, open-label, multicenter, single-arm observational study designed to evaluate changes in fatigue and cognition in MS patients treated with natalizumab. Adults with relapsing MS and initiating natalizumab were enrolled. The primary endpoint was change in Visual Analog Scale for Fatigue (VAS-F) score over 12 weeks. Changes in Modified Fatigue Impact Scale (MFIS) score, Fatigue Severity Scale (FSS) score, and cognitive performance, using Automated Neuropsychological Assessment Metrics (ANAM), were also assessed. Patients (N = 89) had a mean age of 41 years and a median Expanded Disability Status Scale score of 3.0, and 83% had used at least two prior MS therapies. Significant improvements were observed and maintained at 12 weeks in VAS-F (mean ± SD baseline score, 77.7 ± 10.2; mean ± SD change, -14.9 ± 17.1; P < .0001), MFIS (mean baseline score, 59.1 ± 12.2; mean change, -7.4 ± 11.8; P < .0001), and FSS (median baseline score, 6.3 [range, 3.9-7.0]; median change, -0.4 [range, -2.9-1.4]; P < .0001). Cognitive performance remained stable or improved (depending on the ANAM measure). Thus significant improvements in fatigue were maintained over time, and cognitive performance improved or remained stable up to 48 weeks after initiation of natalizumab in MS patients with some degree of fatigue.