medRxiv preprint doi: https://doi.org/10.1101/2021.08.03.21261441; this version posted August 4, 2021. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. It is made available under a CC-BY-NC-ND 4.0 International license . 1 A single intramuscular injection of monoclonal antibody MAD0004J08 induces in healthy 2 adults SARS-CoV-2 neutralising antibody titres exceeding those induced by infection and 3 vaccination 4 Simone Lanini1, Stefano Milleri2, Emanuele Andreano3, Sarah Nosari4, Ida Paciello3, Giulia Piccini5, Alessandra 5 Gentili7, Adhuna Phogat8, Inesa Hyseni5,6, Margherita Leonardi5,6, Alessandro Torelli5, Emanuele Montomoli5,6,10, 6 Andrea Paolini8,9, Andrea Frosini8, Andrea Antinori1, Emanuele Nicastri1, Enrico Girardi1, Maria Maddalena 7 Plazzi1, Giuseppe Ippolito1, Francesco Vaia1, Giovanni Della Cioppa11, Rino Rappuoli3, 12,* 8 9 1Istituto Nazionale per le Malattie Infettive Lazzaro Spallanzani – IRCCS, Rome, Italy 10 2Centro Ricerche Cliniche di Verona, University and Hospital Trust of Verona, Verona, Italy 11 3Monoclonal Antibody Discovery (MAD) Lab, Fondazione Toscana Life Sciences, Siena, Italy 12 4AchilleS Vaccine, Siena, Italy 13 5VisMederi S.r.l, Siena, Italy 14 6VisMederi Research S.r.l, Siena, Italy 15 7CROss Research, Mendrisio, Switzerland 16 8Fondazione Toscana Life Sciences, Siena, Italy 17 9Toscana Life Sciences Sviluppo, Siena, Italy 18 10Department of Molecular and Developmental Medicine, University of Siena, Siena, Italy 19 11Clinical R&D Consultants, Rome, Italy 20 12 Department of Biotechnology, Chemistry and Pharmacy, University of Siena, Siena, Italy 21 22 *Correspondence to Dr. Rino Rappuoli 23 Email: [email protected] 1 NOTE: This preprint reports new research that has not been certified by peer review and should not be used to guide clinical practice. medRxiv preprint doi: https://doi.org/10.1101/2021.08.03.21261441; this version posted August 4, 2021. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. It is made available under a CC-BY-NC-ND 4.0 International license . 24 Summary 25 Background: The emerging threat represented by SARS-CoV-2 variants, demands the development of therapies for 26 better clinical management of COVID-19. MAD0004J08 is an extremely potent Fc-engineered monoclonal antibody 27 (mAb) able to neutralise in vitro all current SARS-CoV-2 variants of concern (VoCs). This ongoing study, evaluates 28 safety, pharmacokinetics and SARS-CoV-2 sera neutralization effect of MAD0004J08 when administered as single 29 dose intramuscularly in healthy adults. 30 31 Method: We conducted a dose escalation study with sequential enrolment of three cohorts, each with an increasing 32 dose level of MAD0004J08 (48mg, 100mg and 400mg). Within each cohort, 10 young healthy adults were 33 randomized with 4:1 ratio to a single intramuscular (i.m.) injection of MAD0004J08 or placebo. The primary 34 endpoint is the proportion of subjects with severe and/or serious treatment emergent adverse events (TEAEs) within 35 7 days post-treatment. Secondary endpoints reported in this paper are the proportion of subjects with solicited 36 TEAEs up 7 days post dosing, MAD0004J08 serum concentrations and neutralising activity versus the original 37 SARS-COV-2 Wuhan virus at different timepoints post-dosing. As post-hoc analyses, we compared the sera 38 neutralising titres of subjects who received MAD0004J08 with those of people that had received the COVID-19 39 BNT162b2 mRNA vaccine in the previous sixty days (n=10) and COVID-19 convalescent patients (n=20), and 40 assessed serum neutralisation activity against the B.1.1.7 (alpha), B.1.351 (beta) and B.1.1.248 (gamma) SARS- 41 CoV-2 variants of concern. 42 43 Findings: A total of 30 subjects, 10 per cohort, were enrolled and randomized. Data up to 30 days were available 44 and analysed in this report. No severe TEAEs were reported in any of the cohorts in the 7 days post-treatment. 45 MAD0004J08 was detected in the sera of treated subjects within few hours post-administration and reached almost 46 maximal levels on day 8. The geometric mean neutralising titres (GMT) assessed against the original Wuhan virus 47 peaked on day 8 and ranged 226 – 905, 905 – 2,560, and 1,280 – 5,120 for cohort 1, 2 and 3 respectively. The sera 48 neutralising GMT in MAD0004J08 treated subjects in all the three cohorts were found to be 1·5- 54·5-fold higher 49 compared to sera from convalescent patients and 1·83– 76·4-fold higher compared to sera from COVID-19 50 vaccinees. Finally, GMT in MAD0004J08 treated subjects showed high neutralising titres versus the B.1.1.7 (alpha), 51 B.1.351 (beta) and B.1.1.248 (gamma) SARS-CoV-2 VoCs. 52 53 Interpretation: A single dose administration of MAD0004J08 via i.m. route is safe and well tolerated and results in 54 a rapid systemic distribution of the MAD0004J08 and sera neutralising titres higher than COVID-19 convalescent 55 and vaccinated subjects. A single dose administration of MAD0004J08 is also sufficient to effectively neutralise 56 major SARS-CoV-2 variants of concern. Based on these results, a Phase 2-3 trial is ongoing to further assess the 57 safety, dosage, and efficacy of MAD0004J08 in asymptomatic or mild-moderate symptomatic COVID-19 patients. 58 59 Funding: EU Malaria Fund, Ministero dello Sviluppo Economico, Ministero della Salute, Regione Toscana, 60 Toscana Life Sciences Sviluppo and European Research Council. 2 medRxiv preprint doi: https://doi.org/10.1101/2021.08.03.21261441; this version posted August 4, 2021. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. It is made available under a CC-BY-NC-ND 4.0 International license . 61 Research in context 62 Evidence before this study 63 We searched PUBMED, MEDLINE and MedRxiv for clinical trials, meta-analyses and randomized controlled trials 64 evaluating the antibody neutralization titres vs. different SARS-CoV-2 variants of concern obtained from subjects 65 who received monoclonal antibodies for the treatment of COVID-19 using the following search terms: (“COVID- 66 19” OR “SARS-CoV-2”) AND (“monoclonal antibody” OR "neutralising antibody") AND ("variants" OR "variants 67 of concern"). No relevant studies were identified. 68 69 Added value of this study 70 This is the first human study assessing safety, PK and neutralising potential of MAD0004J08, a monoclonal 71 antibody against SARS-CoV-2 wild type Wuhan virus and variants of concern, administered intramuscularly at low 72 dosages (48, 100 and 400 mg). MAD0004J08 showed to be safe and well tolerated in the tested dose range. Anti- 73 spike antibodies were detected in the sera of tested SARS-CoV-2 negative healthy adults few hours post-injection. 74 In addition, the sera obtained from MAD0004J08treated subjects, showed to have high neutralisation titres against 75 the Wuhan virus, the B.1.1.7 (alpha), B.1.351 (beta) and B.1.1.248 (gamma) variants of concern. 76 77 Implications of all the available evidence 78 A potent monoclonal antibody such as MAD0004J08, capable of neutralising multiple variants of concern of SARS- 79 CoV-2 rapidly and long lastingly when given as a single intramuscular injection. The antibody, presently tested in a 80 phase 2-3 efficacy trial, can be a major advancement in the prophylaxis and clinical management of COVID-19, 81 because of its broad spectrum, ease of use in non-hospital settings and economic sustainability. 3 medRxiv preprint doi: https://doi.org/10.1101/2021.08.03.21261441; this version posted August 4, 2021. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. It is made available under a CC-BY-NC-ND 4.0 International license . 82 Introduction 83 The COVID-19 pandemic highlighted the potential of human monoclonal antibodies (mAbs) to tackle pandemics as 84 they demonstrated to be safe and effective therapeutic tools that can be brought from discovery to proof-of-concept 85 trials in only 5 – 6 months1. Since the start of the pandemic a dozen mAbs capable of neutralising the SARS-CoV-2 86 virus have been identified and are under clinical development. Regulatory agencies have granted Emergency Use 87 Authorization (EUA) to Eli Lilly’s mAb bamlanivimab (LY-CoV555) and mAb combination bamlanivimab + 88 etesevimab, to Regeneron’s mAb combination casiribimab + imdevimab (REGN-COV2), to Celltrion regdanvimab, 89 and GSK/VIR sotrovimab.2-5 With the emergence of antibody-resistant SARS-CoV-2 variants of concern (VoCs) at 90 the end of 2020, some mAbs lost their clinical efficacy and in April 2021 the FDA revoked EUA for bamlanivimab.6 91 The Toscana Life Sciences Foundation has recently reported the isolation and characterization of the neutralising 92 mAb, MAD0004J08 from a convalescent COVID-19 patient.7 MAD0004J08 appears to be the best candidate for 93 clinical development as it displays the most desirable characteristics for the development of mAb-based prophylaxis 94 and therapy: 1) unprecedented high neutralising potency, implying low dose requirement, intramuscular route of 95 administration, and significant cost reduction of cost of goods. 2) breadth of neutralising activity, in vitro vs. the 96 original Wuhan virus and all current SARS-CoV-2 VoC; B.1.1.7 (alpha), B.1.351 (beta), B.1.1.248 (gamma) and 97 B.1.617.2 (delta) with a potency below 10 ng/mL (7 and manuscript in preparation), implying broad clinical 98 usefulness in countries with diverse SARS-CoV-2 epidemiology.