key: cord-0898130-9nxmwr8d authors: Gobeil, P.; Pillet, S.; Seguin, A.; Boulay, I.; Mahmood, A.; Vinh, D. C.; Charland, N.; Boutet, P.; Roman, F. P.; Van Der Most, R.; de los Angeles Ceregido Perez, M.; Ward, B. J.; Landry, N. title: Interim Report of a Phase 2 Randomized Trial of a Plant-Produced Virus-Like Particle Vaccine for Covid-19 in Healthy Adults Aged 18-64 and Older Adults Aged 65 and Older date: 2021-05-17 journal: nan DOI: 10.1101/2021.05.14.21257248 sha: a2243e7fcb749b691930395aeb518f034c8392dd doc_id: 898130 cord_uid: 9nxmwr8d The rapid spread of SARS-CoV-2 globally continues to impact humanity on a global scale with rising morbidity and mortality. Despite the development of multiple effective vaccines, new vaccines continue to be required to supply ongoing demand. We report Day 42 interim safety and immunogenicity data from a Phase 2, randomized, placebo-controlled trial in Adults aged 18+ immunized with a virus-like particle vaccine candidate produced in plants displaying SARS-CoV-2 spike glycoprotein (CoVLP) adjuvanted with AS03 (NCT04636697). This report focuses on presenting safety, tolerability and immunogenicity, as measured by neutralizing antibody (NAb) and cell mediated immunity (IFN-{gamma} and IL-4 ELISpot) responses, in Adults aged 18-64 (Adults) and Older Adults aged 65+ (Older Adults). CoVLP adjuvanted with AS03 was well-tolerated and adverse events (AE) were primarily mild or moderate and of transient duration. AEs in Older Adults were more limited than those observed in the Adult population. CoVLP with AS03 induced a significant humoral immune response in both age cohorts. CoVLP with AS03 induced a greater humoral response in Adults than Older Adults after a single dose but this effect was overcome with a second dose when both age cohorts responded with NAb titers that were ~10-fold higher than those in a panel of sera from patients recovering from COVID-19. A single dose of CoVLP with AS03 induced a significant IFN-{gamma} response in both age cohorts; a second dose significantly boosted IFN-{gamma} and IL-4 responses in both age cohorts. Adults generated a stronger IFN-{gamma} and IL-4 cellular response than did Older Adults after one or two doses of AS03-adjuvanted CoVLP. Safety and immunogenicity from Adults with comorbidities as well as final safety and immunogenicity responses after 12 months will be reported upon availability. Safety and tolerability data after the first and second doses (as of April 2, 2021) are provided 104 for 306 and 301 subjects in the Adult and for 282 and 272 in the Older Adult groups 105 respectively. Overall, the vaccine was well-tolerated in both populations, with a slightly milder 106 reactogenicity profile in the Older Adults. 107 Reactogenicity is illustrated in Figure 2 for solicited A) local and B) systemic AE. Frequency 108 of solicited AE increased after the second dose relative to the first dose in both age cohorts 109 although Older Adults generally had more muted responses than Adults: 94.5% and 88.4% of 110 Adults reported at least one local AE after the first and second doses respectively compared to 111 61.7% and 54.2% of the Older Adults. In both study populations after the second dose, pain at 112 the injection site was the most frequently reported local AE (89.3% of Adults and 72.8% of 113 Older Adults) while fatigue (67.6% of Adults and 46.1% of Older Adults) and muscle aches 114 (66.0% of Adults and 43.0% of Older Adults) were the most frequently reported systemic AEs. 115 In both populations, the majority of AEs were mild (Grade 1) or moderate (Grade 2) in severity 116 (2.1% and 6.3% of Adults experienced grade 3 AEs after the first and second doses respectively 117 while in Older Adults 0% and 8.7% of participants experienced grade 3 AEs after the first and 118 The seroconversion rates were also comparable between the two age cohorts at D42 (99.2%, 136 95CI: 97.0-99.9 in Adults and 97.7%, 95CI: 93.4-99.5 in Older Adults, p=0.348). 137 Consistent with observations from the Phase 1 study, the NAb titers elicited by CoVLP+AS03 138 at D42 were approximately 10-fold higher than those observed in a panel of convalescent sera 139 (10.2x in Adults and 9.8x in Older Adults). 140 To assist in standardizing the NAb results, the WHO pooled plasma reference standard 20/136, 141 was included in the pseudovirion NAb assay yielding a reference GMT value of 1872. 142 Expressing the GMT results in International Units (IU/mL), the Adults in our Phase 2 study had 143 NAb values of 23.6 and 1083 IU/mL and the Older Adults had values of 16.6 and 1045 IU/mL 144 after the 1 st and 2 nd doses respectively. 145 Prior to vaccination, 9 (1.5%) of the subjects the Phase 2 portion of the study were seropositive 146 at Baseline (6 Adult and 3 Older Adults); all received CoVLP+AS03. At D21, their NAb GMTs 147 were significantly increased in all groups (5756 at D21 relative to 63.9 at D0 in Adults and 148 4909 at D21 relative to 49.6 at D0 in Older Adults). A second dose was only moderately 149 effective in further increasing NAb titers in Older Adults (5786 at D42 in Adults and 10400 at 150 D42 in Older Adults). These data illustrate the potential for CoVLP+AS03 to boost the NAb 151 response even in those presumed to have had prior infection. 152 Overall, CoVLP +AS03 induced comparable NAb responses in both age cohorts. Although 153 small differences between the groups were observed at D21 after the first dose, these 154 differences were overcome by the second dose. These interim data reflect a data cut-off of April 16 th , 2021. 160 As observed in the Phase 1 study of CoVLP+AS03 21 and previously reported in several studies 161 31,32 , a significant minority (~20%) of individuals were observed to have pre-existing (D0) IFN-162 γ responses to the spike protein. There was no relationship between NAb seropositivity and 163 All rights reserved. No reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted May 17, 2021. ; https://doi.org/10.1101/2021.05.14.21257248 doi: medRxiv preprint IFN-γ responses at baseline. In both age cohorts, vaccination with CoVLP+AS03 induced a 164 significant increase in IFN-γ response at D21 which was further significantly increased after a 165 second dose at D42 ( Figure 3A ). Adults had a significantly greater IFN-γ response relative to 166 Older Adults at both D21 (p <0.0001) and D42 (p <0.0001). Sixty-nine percent of the Adults 167 and 51% of Older Adults displayed a detectable IFN-γ response after one dose (D21). The 168 frequencies of responders eliciting a detectable response increased to 96% and 86% after the 169 second dose in Adults and Older Adults respectively. 170 Also consistent with the results from the Phase 1 study, no subject in either group had a 171 measurable IL-4 response pre-vaccination (D0). While a limited portion of subjects responded 172 to CoVLP+AS03 with a detectable IL-4 response after one dose (35% in Adults and 17% in 173 Older Adults), the proportion of 'responders' increased to 94% and 75% after the second dose 174 of CoVLP+AS03 in Adults and Older Adults respectively. Although the second dose of 175 CoVLP+AS03 significantly increased the response in both populations, the Adults had median 176 counts significantly higher than Older Adults (p=0.0002). 177 The Phase 2 portion of the ongoing Phase 2/3 study of CoVLP+AS03 was designed to confirm 180 the selection of CoVLP dose and adjuvant identified in the Phase 1 trial and to assess the 181 performance of the chosen formulation in both adults aged 65 or less in good health, adults 65 182 years and more, and adults with comorbid conditions that put them at increased risk from 183 COVID-19. This interim report focuses on healthy individuals ≥18 years of age. The data for 184 individuals with comorbidities are being compiled and will be released in a separate 185 manuscript. The primary outcomes for the Phase 2 portion of this study focused on short-term 186 (up to D42) safety and tolerability of CoVLP+AS03 and the ability of this novel candidate to 187 induce both NAb and cellular responses to SARS-COV-2 spike protein. Reactogenicity in Adults in the Phase 2 study confirmed the profile observed in the much 189 smaller Phase 1 study in 18-55-year-old adults that received CoVLP(3.75µg) with AS03 190 (n=20). Local reactogenicity was characterized by injection site pain in most subjects while 191 systemically, reactogenicity was characterized by muscle aches, fatigue, and/or headache in 192 approximately 60% of the study participants. There was a trend towards increased local and 193 systemic reactogenicity after the second dose, although almost all reported AEs were mild-to-194 All rights reserved. No reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. was higher in adjuvanted groups as compared to non-adjuvanted groups, as would be expected 198 from its use with other antigens 25,33,34 . As expected 26 and has been observed for mRNA, Phase 1 study 21 as well as extending these observations to older adults. After two doses of 207 CoVLP+AS03, seroconversion occurred in >99% of the subjects and NAb titers were ~10x 208 higher than those observed in convalescent sera. Although the NAb response after the first dose 209 of CoVLP+AS03 was slightly weaker in the Older Adult population than in the Adults, the 210 differences in seroconversion rate and GMTs between the two age cohorts disappeared after the 211 second dose. This observation is consistent with the generally reduced capacity of older 212 individuals to respond to vaccination and with findings for other SARS-CoV-2 vaccines 42-44 . 213 The decreased ability of even healthy older individuals to mount strong immune responses after 214 vaccination is likely multi-factorial including a general decline in immune function (ie: 215 immunosenescence) and chronic low-level inflammation (so-called 'inflammaging) 45 . 216 Consistent with the observations that adjuvants can enhance vaccine-induced responses in older 217 individuals 46 , these results suggest that two-doses of CoVLP+AS03 can overcome these age-218 associated limitations for NAb production at least. 219 The number of Adult or Older Adult subjects in the current study with pre-existing NAb titers 220 to SARS-CoV-2 was very low (n=9, 1.5%) but vaccination with CoVLP+AS03 nonetheless 221 appeared to induce a substantial increase in NAb titers, suggesting that this vaccine candidate 222 can significantly boost a pre-existing memory response 47,48 . This is consistent with 223 observations made by Goel et al. 49 following mRNA vaccination. The increase in NAb titers 224 was greater after the first dose of CoVLP+AS03 (GMT 58.7 → 5458) than the second dose 225 (GMT of 5458 → 7035) suggesting there may be limited additional benefit in providing a 226 All rights reserved. No reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. Although attention on vaccine-induced immune responses for SARS-COV-2 has focused 237 primarily on antibody production, there is compelling evidence that cellular responses 238 contribute to both recovery from infection and long-term immunity 14,32 . Despite this growing 239 consensus, what constitutes a 'good' or 'desirable' response is not yet fully understood 51 . In 240 the current study, the mixed Th1 (IFN-γ) and Th2 (IL-4) cellular response to CoVLP+AS03 241 was, like the NAb response, entirely consistent with the Phase 1 results. In both age cohorts, an 242 IFN-γ dominated response was observed after the first dose that shifted to include a substantial 243 IL-4 response after the second dose. Even though the IL-4 response increased significantly after 244 the second dose, SFU counts for IFN-γ remained approximately 2-fold higher than those for IL-245 4. Of course, the limited nature of using only these two ELISpots to assess the pattern of 246 cellular immunity generated by CoVLP+AS03 should be acknowledged. 247 While Th2-type responses and possible VAED were initially a point of concern in COVID-19 248 vaccine development 52,53 , there has since been no evidence of disease enhancement in either 249 animal models or across all of the human trials reported to date, nor any evidence that Th2 250 responses are associated with VAED 51 . Indeed, it even possible that the Th2-response induced 251 by CoVLP+AS03 as characterized by IL-4 production, may contribute significantly to the high 252 titers of neutralizing antibody observed in vaccinated subjects through its role in T helper 253 follicular cell involvement, optimal germinal center formation, and B-cell maturation 18,54,55 . 254 Such Th2-driven effects may also contribute to longevity of SARS-CoV-2 specific memory B-255 cell responses and longer-term immune response 56 . 256 Unlike the NAb response, both IFN-γ and IL-4 responses in the Older Adults were significantly 257 weaker than those observed in the Adult population even after the second dose suggesting that 258 All rights reserved. No reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted May 17, 2021. ; https://doi.org/10.1101/2021.05.14.21257248 doi: medRxiv preprint at least some aspects of the aged immune system cannot be overcome with multiple doses of an 259 AS03-adjuvanted vaccine. Given the clear age-related differences in both the clinical 260 manifestations of COVID-19 and the immune response generated by SARS-COV-2 infection 261 57 , it is not surprising that vaccine-induced responses might also differ between younger and 262 older individuals 58 . Indeed, similar age-related differences in immune responses have been 263 reported for several of the SARS-COV-2 vaccines in development or in use 38,59 and age-related 264 differences in vaccine efficacy with some of the deployed vaccines are emerging 60-62 . Given 265 the very high NAb titers induced by CoVLP+AS03 across all ages, it is unclear whether or not 266 the in vitro differences in cellular responses between younger and older adults will result in 267 clinically relevant differences in protection. Any such differences might only become apparent 268 as the Phase 3 portion of the study progresses and/or with greater time should CoVLP+AS03 be 269 This study as presented has several obvious limitations. First, the cut-off for data in this 271 interim report was D42 so longer-term safety and immunogenicity results are not yet available. 272 Also, the report does not include Adults with Comorbidities from the Phase 2 study as these 273 data are still being analyzed. Some immunogenicity data, particular for D42 cell-mediated 274 immune responses, have not yet been fully analyzed. Whether or not the NAb induced by 275 CoVLP+AS03 have activity against variants of concern is of obvious interest and these studies 276 are underway. Finally, the limited demographic diversity of the Phase 2 study participants is 277 acknowledged and is primarily a reflection of the demographics of the study sites where the 278 majority of these participants were recruited. The ongoing global Phase 3 efficacy portion of 279 the trial is being conducted on three continents and is expected to provide efficacy results in a 280 highly diverse population. 281 In conclusion, this interim report of our on-going Phase 2/3 study of CoVLP+AS03 confirms 282 that this formulation is well-tolerated and highly immunogenic in healthy adults ≥ 18 years of 283 age. Compared to either a panel of convalescent serum/plasma or the WHO standard serum 284 reagent (20/136), the NAb response induced by CoVLP+AS03 was among the highest reported 285 for any SARS-COV-2 vaccine. Across the broad age range of study participants (18 -88 years 286 of age), >99% mounted either a strong NAb response, a balanced Th1/Th2-pattern cellular 287 response, or both following two doses of CoVLP+AS03. 288 289 290 All rights reserved. No reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. All rights reserved. No reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. All rights reserved. No reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted May 17, 2021. ; https://doi.org/10.1101/2021.05.14.21257248 doi: medRxiv preprint The study was sponsored by Medicago Inc. 479 480 Medicago Inc. is committed to providing access to anonymized data collected during the trial 482 that underlie the results reported in this article, at the end of the clinical trial, which is currently 483 scheduled to be 1 year after the last participant is enrolled, unless granted an extension. 484 Medicago Inc. will collaborate with its partners (GlaxoSmithKline, Rixensart, Belgium) on 485 such requests before disclosure. Proposals should be directed to wardb@medicago.com or 486 landryn@medicago.com. To gain access, data requestors will need to sign a data access 487 agreement and access will be granted for non-commercial research purposes only. The 488 All rights reserved. No reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. Text (7746 words) 500 Tables (1) 502 All rights reserved. No reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted May 17, 2021. ; https://doi.org/10.1101/2021.05.14.21257248 doi: medRxiv preprint was misdosed for the second dose administration and received a placebo in error. For more 508 details of subject disposition, see Table 1 . Fever was defined as oral temperature ≥38.0°C. 518 (which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted May 17, 2021. ; https://doi.org/10.1101/2021.05.14.21257248 doi: medRxiv preprint The CoVLP vaccine candidate has previously been described in detail 21 . Briefly, full-length 545 spike protein from SARS-CoV-2 (strain hCoV-19/USA/CA2/2020) incorporating the 546 modifications R667G, R668S, R670S, K971P, and V972P is expressed in Nicotiana 547 benthamiana by transient transfection, resulting in spontaneous trimer formation and VLP 548 assembly and budding. The VLPs are purified and shipped to the vaccination site where it is 549 mixed with AS03 adjuvant prior to injection. 550 AS03 adjuvant, an oil-in-water emulsion containing DL-α-tocopherol (11.69 mg/dose) and 551 squalene (10.86 mg/dose), was supplied by GlaxoSmithKline. 552 CoVLP was available in single-dose vials (0.30 mL) at 15 µg/ml and stored at 2-8°C until 554 shortly before use. The AS03 adjuvant was supplied in multi-dose vials (10 doses/vial) 555 containing DL-α-tocopherol (53.76 mg/mL) and squalene (43.44 mg/mL). Immediately prior 556 to use 0.3 mL of CoVLP and 0.3 mL of AS03 were mixed gently 1:1 volume: volume in the 557 CoVLP vial and a 0.5 mL withdrawn for injection. All injections were administered 558 intramuscularly using a 23g needle in the deltoid. The first and second doses were administered 559 contralaterally when possible. 560 The phase 2 portion of the study is a randomized, observer-blinded, placebo-controlled study 562 with male and female subjects. antibodies was performed at screening using a commercial ELISA that targets the nucleocapsid 573 (N) protein (ElecSys: Roche Diagnostics), both seronegative and seropositive subjects were 574 enrolled. 575 For Population 1 (Adults), subjects had to be 18-64 years of age. For Population 2 (Older 576 Adults), subjects had to be 65 years of age or older and to be non-institutionalized (eg, not 577 living in rehabilitation centers or old-age homes; living in an elderly community was 578 acceptable). 579 For both study population presented, subjects must have had a body mass index less than 30 580 kg/m 2 and be in good general health with no clinically relevant abnormalities that could 581 jeopardize subject safety or interfere with study assessments, as determined by medical history, family (first-degree relatives) history of narcolepsy, xiv) a history of Guillain-Barré Syndrome. 605 All rights reserved. No reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted May 17, 2021. ; https://doi.org/10.1101/2021.05.14.21257248 doi: medRxiv preprint Enrollment into the Phase 2 portion of the study was closed on 25 March 2021. 606 The participants and the personnel collecting the safety information and working in testing 607 laboratories remained blinded to treatment allocation. On Day 0, D21 and D42, serum and 608 peripheral blood mononuclear cells (PBMC) were processed for immune outcomes. All safety 609 information was collected, and all laboratory procedures were carried out by study staff blinded 610 to treatment allocation. There were no major Protocol changes during the conduct of this study 611 prior to the preparation of the current manuscript. 612 The primary objectives of the Phase 2 portion of the study were to assess safety and tolerability 614 and immunogenicity to CoVLP and AS03 at day 0, 21, and 42 post vaccination compared to 615 placebo in Adults and Older Adults. The safety and immunogenicity data collected at later timepoints in this on-going study will be 631 released once study follow-up has been completed. 632 Both passive (diary) and active monitoring of safety signals were performed for the first 42 634 days of the study and will be continued throughout the study. Active monitoring included 635 All rights reserved. No reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted May 17, 2021. ; https://doi.org/10.1101/2021.05.14.21257248 doi: medRxiv preprint telephone contacts with subjects one and eight days (D1, D8) after each vaccination as well as a 636 site visit on D3 after vaccination. Solicited AEs were assessed by the subjects as Grade 1 to 4 637 (mild, moderate, severe, or potentially life-threatening) according to the criteria previously 638 described 21 . Unsolicited AEs, and AEs leading to subject withdrawal were collected up to D21 639 after each vaccination. The following event(s) would pause or halt the study for further review 640 and assessment of the event(s) by the IDMC: i) Any death, ii) Any vaccine-related SAE; iii) 641 Any life-threatening (Grade 4) vaccine-related AE; iv) If 10 % or more of subjects in a single 642 treatment group, experienced the same or similar listed event(s) that could not be clearly 643 attributed to another cause: v) A severe (Grade 3 or higher) vaccine-related AE; vi) A severe 644 (Grade 3 or higher) vaccine-related vital sign(s) abnormality; vii) A severe (Grade 3 or higher) 645 vaccine-related clinical laboratory abnormality. 646 In the event that a pre-defined safety signal was met in any treatment group, at least a transient 647 halt to the study was planned to permit complete evaluation of the reported event(s) and to 648 consult with the IDMC. 649 Subjects will return to the Investigator site on Days 128, 201, and 386 for safety follow-ups and 650 immunogenicity assessments. 651 Safety signals for VAED, hypersensitivity reactions, and potential immune-mediated diseases 652 were monitored as previously described 21 . 653 Full details of the pseudovirus neutralization assay (Nexelis, Quebec, Canada) have previously 655 been described 21 . Briefly, the assay is based on a genetically modified Vesicular Stomatitis 656 Virus (VSV) from which the glycoprotein G was removed, and a luciferase reporter introduced. 657 The modified VSV vector expresses full length SARS-CoV-2 S glycoprotein (NXL137-1 in 658 POG2 containing 2019-nCOV Wuhan-Hu-1; Genebank: MN908947) from which the last 659 nineteen amino acids of the cytoplasmic tail were removed (rVSVΔG-Luc-Spike ΔCT). 660 Pseudovirions are mixed with vaccinee sera and the degree of neutralization quantified using 661 human ACE-2 expressing VERO cells and reduction in luciferase-based luminescence. For 662 each sample, the neutralizing titer was defined as the reciprocal dilution corresponding to the 663 50% neutralization (NT50), when compared to the pseudoparticle control. Samples below cut-664 off were attributed a value of 5 (half the minimum required dilution). 665 All rights reserved. No reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted May 17, 2021. ; https://doi.org/10.1101/2021.05.14.21257248 doi: medRxiv preprint FIGURES 726 (which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted May 17, 2021. ; https://doi.org/10.1101/2021.05.14.21257248 doi: medRxiv preprint (which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted May 17, 2021. ; https://doi.org/10.1101/2021.05.14.21257248 doi: medRxiv preprint 732 All rights reserved. No reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted May 17, 2021. ; https://doi.org/10.1101/2021.05.14.21257248 doi: medRxiv preprint (which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted May 17, 2021. ; https://doi.org/10.1101/2021.05.14.21257248 doi: medRxiv preprint (which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted May 17, 2021. ; https://doi.org/10.1101/2021.05.14.21257248 doi: medRxiv preprint All rights reserved. No reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted May 17, 2021. ; https://doi.org/10.1101/2021.05.14.21257248 doi: medRxiv preprint Follicular helper CD4 T cells (TFH) Immunological memory to SARS-CoV-2 assessed for up to 8 months 439 after infection Age-Related Differences in Immunological Responses to SARS Immune Response With Aging: Immunosenescence and Its Potential Impact on COVID-19 Age-related heterogeneity in immune responses to SARS-CoV-2 vaccine BNT162b2. medRxiv Effectiveness of the Pfizer-BioNTech COVID-19 Vaccine Among 449 Residents of Two Skilled Nursing Facilities Experiencing COVID-19 Outbreaks -Connecticut Efficacy and Safety of the mRNA-1273 SARS-CoV-2 Vaccine Early effectiveness of COVID-19 vaccination with BNT162b2 To facilitate the comparability of results across different trials, the WHO International Standard 666 for anti-SARS-CoV-2 immunoglobulin (human; NIBSC code: 20/136) was established to allow 667 conversion of neutralization assay titers into international units (IU/mL) Upon multiple assessments using this validated PNA assay, a conversion factor of 1.872 was 670 established. Hence, the antibody titers presented throughout this manuscript can be expressed 671 as IU/mL by dividing the NT50 by this factor ELISpot 673 PBMC samples from study subjects were analyzed by IFN-γ or IL-4 ELISpot (Caprion Canada) using a pool of 15-mer peptides with 11aa overlaps from SARS-CoV-2 S 675 protein (USA-CA2/2020, Genbank: MN994468.1, Genscript, purity >90%). Full details of the 676 methodology are detailed elsewhere 21 Convalescent serum and plasma samples Sera/plasma from COVID-19 convalescent patients were collected from a total of 35 679 individuals with confirmed disease diagnosis. Time between the onset of the symptoms and 680 sample collection varied between 27 and 105 days. Four sera samples were collected by ) and 20 sera samples by Sanguine BioSciences Eleven plasma samples were 683 collected from previously hospitalized patients at McGill University Health Centre. Disease 684 severity were ranked as mild (COVID-19 symptoms without shortness of breath), moderate 685 (shortness of breath reported), and severe (hospitalized) Analysis Populations and Statistical Analysis Plan The sample size of 306 691 vaccinated Adults and 282 Older Adults made it possible to perform the initial evaluation of the 692 vaccine immunogenicity and detect major differences in rates of AEs between groups. The 693 sample size was not large enough to detect all types of The 697 ITT consists of all randomized subjects who received the CoVLP and AS03 or placebo and 698 analyzed prior to the data cut (Adults: Day 21 n=299; Day 42 n=299; Older Adults Day 21 699 n=255; Day 42 n=165). Immunogenicity was evaluated by humoral immune response (NtAb 700 assays) and cell-mediated immune (CMI) response (ELISpot) induced in subjects on D0, 21 701 and 42. To assess the humoral immune response, the GMT was calculated and compared 702 between CoVLP and AS03 and placebo using an ANOVA on the log-transformed titers. The 703 log transformation was used to meet the normal assumption for the ANOVA Adults at Day 21 and Day 42 using an ANOVA. Fisher's exact test was used to compare 707 seroconversion rate between the treatment groups. The 95% CI for seroconversion was 708 calculated using the exact Clopper-Pearson method. The specific T helper type 1 (Th1) and Th2 CMI responses along with the corresponding 95% CI for the median induced on D0, D21 and 710 D42 were measured by the number of T cells expressing IFN-γ and IL-4 respectively Since the response between 713 timepoints are paired data, the Wilcoxon Signed Rank Test was used to estimate the difference 714 in response between timepoints, along with the corresponding 95% CI for the median 715 separately for each treatment group. The difference in IFN-γ and IL-4 were also compared 716 between Adults Safety assessment are based on the Safety Analysis Set, i.e. all subjects who received either the CoVLP candidate vaccine with or without an adjuvant. Occurrence and incidence of safety 719 events were reported for each treatment groups. No formal hypothesis-testing analysis of AE 720 incidence rates was performed All authors contributed significantly to the submitted work. BJ Ward, N Landry, A Seguin 461 contributed to all aspects of the clinical study from conception to completion. P Gobeil, A.