Fernando Rodríguez-Jorge ¹ José Ignacio Fernández-Velasco ^2* Noelia Villarrubia ² Julia Gracia-Gil ³ Eva Fernández ³ Virginia Meca-Lallana ⁴ Carolina Díaz-Pérez ⁴ Susana Sainz de la Maza ¹ Eva María Pacheco ⁵ Ana Quiroga ⁶ Lluis Ramió-Torrentà ⁶ Sergio Martínez-Yélamos ⁷ Laura Bau ⁷ Enric Monreal ⁸ Ana López-Real ⁹ Alexander Rodero Romero ² Laura Borrega ¹⁰ Santiago Díaz ¹¹ Pablo Eguía ¹¹ Mercedes Espiño ² Juan Luis Chico García ¹ Francisco Javier Barrero ¹² Marisa Martínez Ginés ¹³ José M. García-Domínguez ¹³ Irene Moreno ¹⁴ Raquel Sainz-Amo ^1,2 M Alba Mañé-Martínez ¹⁵ A. Caminero ¹⁶ Fernando Castellanos ¹⁷ Andrés Labiano-Fontcuberta ¹⁸ Lucía Ayuso ¹⁹ Rossana Abreu ²⁰ Miguel Ángel Hernández ²⁰ José Meca-Lallana ²¹ Lorena Martín-Aguilar ²² Jaime MASJUAN ¹ Lucienne Costa-Frossard ¹ Luisa M. Villar ²

• ¹ Department of Neurology, Ramón y Cajal University Hospital, Madrid, Spain
• ² Department of Immunology, Ramón y Cajal University Hospital, Madrid, Catalonia, Spain
• ³ Department of Neurology, Albacete University General Hospital, Madrid, Spain
• ⁴ Department of Neurology, La Princesa University Hospital, Madrid, Catalonia, Spain
• ⁵ Department of Neurology, Juan Ramón Jiménez University Hospital, Huelva, Spain
• ⁶ Department of Neurology, Doctor Josep Trueta Girona University Hospital, Gerona, Spain
• ⁷ Department of Neurology, Bellvitge University Hospital, Barcelona, Balearic Islands, Spain
• ⁸ Department of Neurology, Ramón y Cajal University Hospital, Madrid, Madrid, Spain
• ⁹ Department of Neurology, A Coruña University Hospital Complex, La Coruña, Spain
• ¹⁰ Department of Neurology, Fundación Alcorcón University Hospital, Madrid, Asturias, Spain
• ¹¹ Department of Neurology, Gran Canaria Doctor Negrín University Hospital, Gran Canaria, Spain
• ¹² Department of Neurology, Clínico San Cecilio University Hospital, Granada, Spain
• ¹³ Department of Neurology, Gregorio Marañón University Hospital, Madrid, Asturias, Spain
• ¹⁴ Department of Neurology, Fundación Jiménez Díaz University Hospital, Madrid, Asturias, Spain
• ¹⁵ Department of Neurology, Joan XXIII University Hospital, Tarragona, Spain
• ¹⁶ Department of Neurology, Ávila Hospital Complex, Ávila, Spain
• ¹⁷ Department of Neurology, Virgen del Puerto Hospital, Cáceres, Spain
• ¹⁸ Department of Neurology, Doce de Octubre University Hospital, Madrid, Spain
• ¹⁹ Department of Neurology, Príncipe de Asturias University Hospital, Madrid, Spain
• ²⁰ Department of Neurology, Nuestra Señora de Candelaria University Hospital, Tenerife, Spain
• ²¹ Department of Neurology, Virgen de la Arrixaca University Hospital, Murcia, Spain
• ²² Department of Neurology, Sant Creu i Sant Pau Hospital, Barcelona, Spain

Objective: To ascertain the changes of serum neurofilament light chain (sNfL) and glial fibrillary acidic protein (sGFAP) values in relapsing-remitting multiple sclerosis (RRMS) patients treated with ocrelizumab and their association with treatment response. Methods: Multicenter prospective study including 115 RRMS patients initiating ocrelizumab treatment between February/2020 and March/2022 followed during a year. Serum samples were collected at baseline and every 3 months to measure sNfL and sGFAP levels using single molecule array (SIMOA) technology. Based on age and body mass index, sNfL values were standardized using z-score. NEDA (Non-evidence of disease activity)-3 status was defined for patients free of disease activity after a year of follow-up. Inflammation (INFL) was considered when new relapses occurred during follow-up, or new MRI lesions were found at one-year exploration. PIRA (progression independent of relapse activity) was defined as disability progression occurring in absence of relapses or new MRI activity. Results: After a year on ocrelizumab, 85 patients (73.9%) achieved NEDA-3. Thirty patients did not achieve NEDA: 20 (17.4%) because of INFL and 10 (8.7%) because of PIRA. Of INFL patients: 6 (30.0%) had relapses, and 17 (85.0%) had at least one new MRI lesion on 12 months examination. At baseline, INFL patients had higher sNfL (p=0.0003) and sGFAP (p=0.03) than NEDA-3 group. PIRA patients mostly exhibited low sNfL and heterogeneous sGFAP levels. After a year, NEDA-3 and INFL patients showed similar decreases in sNfL (p<0.0001) and sGFAP (p<0.0001 for NEDA-3 and p=0.001 for INFL ones). However, the decrease occurred earlier in NEDA-3 patients. Accordingly, sNfL>1.5 z-score three months after ocrelizumab initiation indicated a higher risk of inflammation (OR=13.6; p<0.0001). Decrease of sGFAP values occurred later in both groups, with significant reductions observed at 12 months for INFL and 6 and 12 months for NEDA-3. No significant changes in sNfL or sGFAP were observed in PIRA patients. Conclusion: Ocrelizumab induced normalization of sNfL and sGFAP in the majority of NEDA-3 and inflammatory patients but did not cause changes in PIRA group. Our data suggest that normalization of sNfL and sGFAP associate with the lack of inflammatory-associated disease progression but it may not affect non-inflammatory PIRA.