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Background and purpose: An enhanced severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2) vaccine regimen could improve humoral vaccine response in patients with multiple sclerosis (MS) treated by anti‐CD20. The aim was to evaluate the serological response and the neutralizing activity after BNT162b2 primary and booster vaccination in MS patients, including patients on anti‐CD20 receiving a primary vaccine regimen enhanced with three injections. Methods: In this prospective longitudinal cohort study of 90 patients (47 on anti‐CD20, 10 on fingolimod, 33 on natalizumab, dimethylfumarate or teriflunomide), anti‐SARS‐CoV‐2 receptor binding domain (RBD) immunoglobulin G antibodies were quantified and their neutralization capacity was evaluated by enzyme‐linked immunosorbent assay (GenScript) and a virus neutralization test against B.1 historical strain, Delta and Omicron variants, before and after three to four BNT162b2 injections. Results: After the primary vaccination scheme, the anti‐RBD positivity rate was strongly decreased in patients on anti‐CD20 (28% [15%; 44%] after two shots, 45% [29%; 62%] after three shots) and fingolimod (50% [16%; 84%]) compared to other treatments (100% [90%; 100%]). Neutralization activity was also decreased in patients on anti‐CD20 and fingolimod, and notably low for the Omicron variant in all patients (0%–22%). Delayed booster vaccination was performed in 54 patients, leading to a mild increase of anti‐RBD seropositivity in patients on anti‐CD20 although it was still lower compared to other treatments (65% [43%; 84%] vs. 100% [87%; 100%] respectively). After a booster, Omicron neutralization activity remained low on anti‐CD20 and fingolimod treated patients but was strongly increased in patients on other treatments (91% [72%; 99%]). Discussion: In MS patients on anti‐CD20, an enhanced primary vaccination scheme moderately increased anti‐RBD seropositivity and anti‐RBD antibody titre, but neutralization activity remained modest even after a fourth booster injection. Trial registration information: COVIVAC‐ID, NCT04844489, first patient included on 20 April 2021. [ABSTRACT FROM AUTHOR]