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SUPPLEMENTARY APPENDIX

Induction of Cross-reactive Hemagglutination Inhibiting Antibody and Polyfunctional CD4+ T-cell Responses by a Recombinant Matrix-M-Adjuvanted Hemagglutinin Nanoparticle

Influenza Vaccine

Corresponding Author: Vivek Shinde

TABLE OF CONTENTS

1 METHODS

1.1 Inclusion and exclusion criteria

1.2 Randomization and stratification

1.3 Blinding

1.4 Study objectives

1.5 Recombinant quadrivalent nanoparticle influenza vaccine (qNIV), Matrix-M

adjuvant, and the comparators

1.6 Safety assessments

1.7 Assays for assessment of antibody and cellular immune responses

1.7.1 Antibody responses assessed by wild-type hemagglutination-inhibition

(wt-HAI) assay

1.7.2 Cellular immune responses assessed by CD4+ T-cell responses on

intracellular cytokine staining (ICCS)

1.8 Statistical analysis

1.9 Ethics, trial registration, and funding source

1

2 SUPPLEMENTARY TABLES AND FIGURES

Figure S1. Neutralizing antibody (MN) responses (GMFRs) induced by tNIV or IIV3-HD

against egg-adapted versus wild-type A/Singapore (H3N2) from phase 1/2 study

Table S1. Trial Design, Treatment Assignments, and Enrollment Stages

Table S2. Schedule of Study Procedures

Table S3. Summary of Baseline Demographic Data Through Day 182 – Safety Population

Table S4. Wild-type (wt) HAI Responses by Treatment Group, Strain, and Time Point

Table S5A. Geometric Mean Counts and Geometric Mean Fold Rise of Specific CD4 T-

Cells Expressing Double and Triple Cytokines per 106 CD4 T-Cells – Intent-to-Treat

Population

Table S5B. Baseline Adjusted Ratio of Geometric Mean Counts of Specific CD4 T-Cells

Expressing Double and Triple Cytokines per 106 CD4 T-Cells in Quad-NIV Groups B and C

vs IIV3-HD – Intent-to-Treat Population

Table S5C. Baseline Adjusted Ratio of Geometric Mean Counts of Specific CD4 T-Cells

Expressing Double and Triple Cytokines per 106 CD4 T-Cells in Quad-NIV Groups B and C

vs RIV4 – Intent-to-Treat Population

Table S6. Summary of All and Severe Solicited Adverse Events From Day 0 Post-

vaccination to Day 6, Inclusive – Safety Population

Table S7. Summary of Severe Adverse Events (SAEs) From Day 0 Post-vaccination to Day

182, Inclusive, by System Organ Class and Preferred Term – Safety Population

3 REFERENCES

2

1 METHODS

1.1 Inclusion and exclusion criteria

Subjects met the following criteria to be eligible to participate:

1) Clinically stable adult male or female, ≥65 years of age. Subjects may have 1 or more

chronic medical diagnoses, but should be clinically stable as assessed by:

Absence of changes in medical therapy within 1 month due to treatment failure or

toxicity

Absence of medical events qualifying as serious adverse events within 2 months

Absence of known, current, and life-limiting diagnoses which render survival to

completion of the protocol unlikely in the opinion of the investigator

2) Willing and able to give informed consent prior to trial enrollment, and living in the

community and able to attend trial visits, comply with trial requirements, and provide

timely, reliable, and complete reports of adverse events.

Subjects were excluded if they met any of the following criteria:

1) Participation in research involving investigational product (drug/biologic/device) within 45

days before planned date of first injection.

2) Participation in any of Novavax’s previous influenza vaccine clinical trial(s).

3) History of a serious reaction to prior influenza vaccination, known allergy to constituents of

Fluzone® High-Dose, Flublok® Quadrivalent, or polysorbate 80.

4) History of Guillain-Barré syndrome within 6 weeks following a previous influenza vaccine.

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5) Received any vaccine in the 4 weeks preceding the trial vaccination and any influenza

vaccine within 6 months preceding the trial vaccination.

6) Any known or suspected immunosuppressive illness, congenital or acquired, based on

medical history and/or physical examination.

7) Chronic administration (defined as more than 14 continuous days) of immunosuppressants

or other immune-modifying drugs within 6 months prior to the administration of the trial

vaccine. An immunosuppressant dose of glucocorticoid will be defined as a systemic dose

≥10 mg of prednisone per day or equivalent. The use of topical, inhaled, and nasal

glucocorticoids will be permitted.

8) Administration of immunoglobulins and/or any blood products within the 3 months

preceding the administration of the trial vaccine or during the trial.

9) Acute disease at the time of enrollment (defined as the presence of a moderate or severe

illness with or without fever, or an oral temperature ≥38.0°C, on the planned day of vaccine

administration).

10) Any condition that in the opinion of the investigator would pose a health risk to the subject

if enrolled or could interfere with evaluation of the vaccine or interpretation of trial results

(including neurologic or psychiatric conditions deemed likely to impair the quality of safety

reporting).

11) Known disturbance of coagulation.

12) Suspicion or recent history (within 1 year of planned vaccination) of alcohol or other

substance abuse.

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1.2 Randomization and stratification

Participant randomization was conducted using an interactive web randomization system.

Stratification was based on age (60 to <75 and ≥75 years), gender, and history of receipt of

2016-2017 influenza vaccine.

1.3 Blinding

Subjects were assigned a subject ID by use of the contract research organization electronic data

capture system. The unblinded vaccine administrator(s) referred to the randomization schedule

for the correct treatment assigned to each subject. All treatment assignments were checked

and confirmed by the assisting unblinded staff. Study personnel preparing and/or administering

the investigational products had no involvement in monitoring subject safety or tolerability, or

in interpreting the study data. Since the physical appearance of the placebo was different from

the vaccine, study subjects were not allowed to view the injection solution.

1.4 Study objectives

The primary objectives were: 1) to describe the safety and tolerability of each test vaccine, and

2) to demonstrate a Matrix-M adjuvant effect by demonstrating the immunogenic superiority of

the quadrivalent nanoparticle influenza vaccine (qNIV) (60 µg hemagglutinin [HA] per A and B

strain) co-formulated with 50 µg Matrix-M1, as compared to qNIV (60 µg HA per A and B strain)

without adjuvant, based on Day 28 post-vaccination wild-type hemagglutination-inhibition (wt-

HAI) antibody responses against 4 vaccine-homologous (2 influenza A and 2 influenza B strains),

and/or 2 antigenically drifted A/H3N2 strains. Secondary or exploratory objectives included: 1)

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to describe the immunogenicity of various formulations of qNIV (Groups A, B, C, D) relative to

the 2 comparators IIV3-HD, or RIV4, based on Day 0 and 28 wt-HAI antibody responses to

vaccine-homologous and antigenically drifted A/H3N2 strains; 2) to describe the longevity of

immune responses of each test vaccine at various time points post-vaccination based on wt-HAI

antibody responses against vaccine-homologous and antigenically drifted strains; 3) to describe

(in a random subset of subjects) the immunogenicity of various formulations of qNIV (Groups A,

B, C, D) relative to the 2 comparators, IIV3-HD or RIV4, based on Day 0, 28, and 56 neutralizing

antibody (MN) responses to vaccine-homologous and antigenically drifted A/H3N2 strains (to

be described in a separate report); and 4) to describe the quality and amplitude of cell-

mediated immune (CMI) responses in a subset of all subjects from 3 pre-selected study sites,

based on functional antigen-specific CD4+ T-cell responses on intracellular cytokine staining

(ICCS) analysis at Days 0 and 7.

1.5 Recombinant quadrivalent nanoparticle influenza vaccine (qNIV), Matrix-M adjuvant, and

the comparators

qNIV was manufactured as previously described and consists of a purified, recombinant full-

length HA that self-assembles into distinct nanoparticle structures of approximately 20 to 40

nm [1]. The baculovirus/Sf9 insect cell system was used to express recombinant influenza HAs.

In Sf9 insect cells, recombinant, uncleaved HAs are produced that are glycosylated and

assemble into homotrimers. Purified, HA homotrimers form higher-order structures composed

of 2 to 9 or more trimers per nanoparticle based on hydrophobic interactions. qNIV was

constructed using wt virus HA gene sequences for A/Hong Kong/4801/14 (H3N2),

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A/Michigan/45/15 (H1N1), and B/Brisbane/60/08 accession numbers EPI834581, EPI699812,

EPI394895, respectively (GISAID Epiflu database).

The qNIV was formulated at 2 HA protein dose levels [either 60 µg per strain for each A and B

strain for a total of 240 µg of HA [Groups A, B, C, E] or 60 µg per A strain and 90 µg per strain for

a total of 300 µg of HA [Group D]), in a 25 mM sodium phosphate buffer, pH 7.5, with 150 mM

sodium chloride, 100 mM arginine hydrochloride, 5% w/v trehalose, and 0.03% w/v polysorbate

80. Matrix-M™, a saponin-based adjuvant, was manufactured as previously described, and

formulated in a phosphate buffer, pH 7.2, with 150 mM sodium chloride [2]. The qNIV antigens

were co-administered with Matrix-M using either in-clinic contemporaneous mixing (Group A)

or co-formulation (Groups B, C, and D) strategies. Co-formulated treatments were mixed and

vialed at least 3 months prior to administration and characterized for stability of both antigens

and adjuvant.

The first licensed comparator vaccine, a trivalent high-dose split inactivated influenza vaccine

(IIV3-HD), was Fluzone® High-Dose (Sanofi Pasteur, Swiftwater, PA), and according to

manufacturer’s product label, contained 180 µg (60 µg per strain) total HA content [3].

The second licensed comparator vaccine, a quadrivalent recombinant HA influenza vaccine

(RIV4), was Flublok® Quadrivalent (Protein Sciences Corporation, Meriden, CT [distributed by

Sanofi Pasteur, Swiftwater, PA]), and according to manufacturer’s product label, contained 180

µg (45 µg per strain) total HA content [4].

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On trial Day 28, participants in Group E were administered a rescue injection with a licensed

seasonal influenza vaccine (IIV3-HD; Fluzone High-Dose), given that the unadjuvanted

formulation of qNIV was being evaluated for the first time in this trial. To maintain the trial

blinding, sterile saline was used as placebo for subjects in Groups A, B, C, D, F, and G for their

Day 28 injection (supplied in a single-dose vial at a concentration of 9 mg/mL in a 2 mL volume

by VWR International, Radnor, PA).

All investigational vaccines contained the World Health Organization (WHO) recommended

quadrivalent or trivalent influenza virus strains for the 2018-2019 Northern Hemisphere

influenza season: A/Michigan/45/2015(H1N1)-like (clade 6B.1); A/ Singapore/INFIMH-16-

0019/2016 (H3N2)-like (clade 3C.2a1); B/Colorado/60/2017 (Victoria-lineage)-like (clade

V1A.1); and B/Phuket/3073/2013 (Yamagata-lineage)-like (clade Y3).19 [5].

1.6 Safety assessments

Staged enrollment to monitor for safety

Enrollment was divided into 3 stages to monitor safety. Stage 1 enrolled a total of 120 subjects

(approximately 20 subjects per treatment group, excluding Group D for which no subjects were

enrolled in Stage 1). Stage 2 enrolled a total of 220 subjects (approximately 20 [Groups A, C, D,

F, and G] or 60 subjects [Groups B and E] per treatment group; Table S1). The remainder of the

subjects (1035 subjects total, that is, 115 [Groups A, C, D, F, and G] and 230 subjects [Groups B

and E] per treatment group; Figure 1) were enrolled in Stage 3. Progression from Stage 1 to

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Stage 2 and from Stage 2 to Stage 3 required favorable review of safety data from the prior

stage against pre-specified vaccination holding rules.

1.7 Assays for assessment of antibody and cellular immune responses

1.7.1 Antibody responses assessed by wild-type hemagglutination-inhibition

(wt-HAI) assay

Due to the documented inability of recent A(H3N2) strains to agglutinate avian or small

mammal red blood cell (RBC) reagents in hemagglutination-inhibition (HAI) assays and, in

addition, the presence of immunologically significant mutations induced by egg passage in the

HA of these strains, vaccine immunogenicity was assessed by the classical HAI qualified method

adapted with 140-190 nm recombinant wild-type hemagglutinin virus-like particles (wt-

HA-VLPs), reflecting the amino acid sequence of circulating virus, as the agglutinating agent and

human type-O RBCs(Biological Specialty Corporation, Colmar, PA) as the agglutination target in

order to restore assessment of HAI antibody activity [6, 7]. Wt-HA-VLPs were constructed with

wild-type HA, neuraminidase (NA), and M1 genes and produced in baculovirus/Sf9 cell

expression system (using HA antigens described in section 1.5).

Briefly, Day 0, 28, 56, and 182 human sera treated with receptor destroying enzyme (RDE;

Denka Seiken Co., Campbell, CA) overnight at 37⁰C to remove non-specific inhibitors of

hemagglutination, heat inactivated at 56⁰C for 30 min, diluted to 1:10 with Dulbecco's

phosphate-buffered saline, plated into microtiter wells, followed by a series of 2-fold dilutions

of serum for a total of 10 dilutions. Four HA units of HA-VLPs in dilution buffer containing 80 nM

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oseltamivir and indicator 0.75% human RBC suspension were added to designated wells in 2

sequential steps, with mixing and incubation at each step in round bottom 96-well plates. Wt-

HAI antibody titers were determined as the reciprocal of the highest dilution of serum that

completely inhibited HA. The serum wt-HAI titer was calculated from the geometric mean titers

(GMTs) of duplicate test results. Positive and negative serum samples were included in each

assay run to serve as assay quality controls. Plates were read on Cypher One Haemagglutination

Analyzer (InDevR Inc., Boulder, CO) to capture plate images.

1.7.2 Cellular immune responses assessed by CD4+ T-cell responses on intracellular cytokine

staining (ICCS)

Human peripheral blood mononuclear cells (PBMCs) were cultured overnight after thawing.

Cells with a viability >75% proceeded to the following assays. Human PBMCs were cultured in

96-well U-bottom plates at a density of 1 x 106 cells/well and treated with the HA protein

(A/Singapore, A/Michigan, A/Wisconsin, or B/Colorado; 5 µg/mL), PMA + ionomycin (a positive

control for T-cell activation), or medium only (negative control). After incubation at 37°C for 6

hours in the presence of BD GolgiPlug™ and BD GolgiStop™ (BD Biosciences, San Jose, CA) for

the last 4 hours of culturing, cells were labeled for surface markers (CD3, CD4, CD8, CD45RA,

and CCR7 [BD Biosciences, San Jose, CA]) and the LIVE/DEAD indicator dye (Life Technologies,

a brand of Thermo Fisher Scientific, Waltham, MA) was added. The intracellular cytokines were

detected by antibodies specific for interferon gamma (IFN-γ), tumor necrosis factor alpha (TNF-

α), and interleukin-2 (IL-2) [BD Biosciences, San Jose, CA]. The samples were processed using a

BD LSR-Fortessa flow cytometer (Becton Dickinson and Company, San Jose, CA). Data were

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analyzed using Flowjo software version Xv10 (Tree Star Inc., Ashland, OR). Data shown were

gated on effector memory CD4+ T-cell populations (CD45RA-CCR7-).

1.8 Statistical analysis

The safety population, defined as all participants who provided consent, were randomized, and

received any investigational treatment, was used for all descriptive safety analyses. The

immunogenicity per protocol (iPP) population, defined as all participants in the safety

population who received the assigned investigational treatment according to the protocol, had

wt-HAI serology results for Days 0 and 28, and had no major protocol deviations affecting the

primary immunogenicity outcomes as determined by the sponsor prior to database lock, and

unblinding was used for all wt-HAI immunogenicity analyses. The intent-to-treat (ITT)

population did not differ meaningfully from the iPP population for purposes of wt-HAI

immunogenicity analyses and is not discussed further here except as follows for CMI. The ITT

population was used to describe all exploratory CMI analyses since iPP criteria for serology

were not relevant to collection and testing of PBMCs for purposes of CMI analyses at Days 0

and 7.

Demographic parameters and other baseline characteristics were summarized by treatment

group for all subjects in the safety population. Continuous variables were presented by

summary statistics (eg, mean and standard deviation [SD] for the non-immunogenicity

endpoints; geometric means and their 95% confidence intervals (CIs) for the immunogenicity

endpoints). Categorical variables were presented by frequency distributions (frequency counts

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and percentages for the non-immunogenicity endpoints; percentages and their 95% CIs for the

immunogenicity endpoints).

Data concerning wt-HAI titers were expressed as GMTs, defined as the antilog of the mean of

the log-transformed wt-HAI titers at a given time point, geometric mean fold rise (GMFRPost/Pre),

defined as the ratio of the Day 28 (post-vaccination) GMTs to Day 0 (pre-vaccination) GMTs,

seroconversion rate (SCR), defined as the percentage of subjects with either a baseline (Day 0)

titer <1:10 and a post-vaccination titer ≥1:40, or a baseline (Day 0) titer ≥1:10 and a post-

vaccination titer ≥4-fold higher at a given post-vaccination time point, and seroprotection rate

(SPR), defined as the proportion of subjects with a reciprocal HAI titer ≥40 at a given time point.

Analyses of wt-HAI GMTs and GMFRs were performed using log normal distribution, and the

antilog of the means or mean treatment group differences along with the corresponding CIs of

the log-transformed titers. Analyses of SCRs were performed using the Clopper-Pearson

method for CIs for each treatment group and the Newcombe method for CIs for the treatment

group differences. Comparisons of wt-HAI antibody responses between the 2 treatments were

calculated as the ratio of GMTs (GMTR) between treatment arms at Day 28 (adjusted for

intergroup variation in the baseline Day 0 titers) using a linear mixed model with baseline titers

included as a covariate in the model to calculate between-group differences in a log10 scale.

Titer values recorded as below the assay lower limit of quantitation (LLOQ) (ie, below the

starting dilution of the assay reported as “<10”) of the assay were set to half LLOQ (ie, 10 / 2 =

5) for the purposes of GMT and GMR analyses.

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For CMI responses measured by ICCS, peripheral blood CD4+ T-cell numbers producing IL-2,

IFN-γ, and TNF-α cytokines following in vitro re-stimulation with influenza vaccine-homologous

(A/Singapore; A/Michigan [H1N1]; B/Colorado [Victoria]) or drift (A/Wisconsin [H3N2]) strain-

specific HAs were reported as absolute median counts (and associated interquartile ranges) and

geometric mean counts (GMCs; and associated 95% CI) of double- or triple-cytokine producing

influenza strain-specific CD4+ T-cells (for each strain), and corresponding geometric mean count

fold rise (GMFRPost/Pre), defined as the ratio of the Day 7 (post-vaccination) GMCs to Day 0 (pre-

vaccination) GMCs (and associated 95% CI). Between-group differences were reported as the

ratio of GMCs (GMCR) between treatment arms at Day 7 (adjusted for intergroup variation in

baseline Day 0 GMCs) of double- or triple-cytokine producing influenza strain-specific CD4+ T

cells.

The primary immunogenicity objective of demonstrating an adjuvant effect required

establishing immunogenic superiority of Group B (Matrix-M adjuvanted qNIV) relative to Group

D (unadjuvanted qNIV) by excluding values ≤1.0 at the lower 95%

CI bound for the baseline-adjusted ratio of Day 28 post-vaccination wt-HAI titers (ie, GMT of

Group B [adjuvant]/GMT of Group E [no adjuvant]) for not less than 2 out of 6 influenza strains

(ie, any 2 of 4 vaccine-homologous strains and/or 2 antigenically drifted influenza strains), while

no other strain(s) demonstrated baseline-adjusted ratios of Day 28 post-immunization HAI

titers that were significantly <1.0.

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A total sample size of 1350 was selected to achieve the primary immunogenicity objective of

demonstrating an adjuvant effect. This study was designed to provide ≥80% power for

demonstrating immunogenic superiority (Group B [adjuvanted qNIV] vs Group E [unadjuvanted

qNIV]) against 2 strains assuming a true difference of 1.5-fold, and non-inferiority against 4

strains assuming a true difference of 1.0-fold.

All other immunogenicity and safety analyses were descriptive, with the purpose of

determining a final dose and formulation for the next study. All statistical analyses (calculations

of CIs and P-values) were performed without formal adjustment for multiple comparisons. For

safety endpoints, the probability of observing at least 1 adverse event among 135 (Group D),

155 (Groups A, C, F, and G), or 310 (Groups B and E) subjects for each of the treatments groups

was >90% if the true rate of such events was 1.7%, 1.5%, or 0.8%, respectively.

All statistical analyses were performed in SAS statistical software (Version 9.4).

1.9 Ethics, trial registration, and funding source

All trial participants provided written informed consent. The trial was approved by an

institutional review board, Quorum Review, Seattle, WA, USA, and conducted in accordance

with the Declaration of Helsinki and International Conference on Harmonisation Guidelines for

Good Clinical Practice. The study was funded by the sponsor, Novavax (Gaithersburg, MD, USA),

and registered in ClinicalTrials.gov (NCT03658629).

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2 SUPPLEMENTARY TABLES AND FIGURES

Figure S1. Neutralizing antibody (MN) responses (GMFRs) induced by tNIV or IIV3-HD against

egg-adapted versus wild-type A/Singapore (H3N2) from phase 1/2 study.

[TIFF figure to replace figure in Word document]

Abbreviations: CI, confidence interval; GMFR, geometric mean fold rise; IIV3-HD, trivalent high-dose inactivated influenza

vaccine; tNIV, trivalent nanoparticle influenza vaccine.

Full strain name: A/Singapore/INFIMH-16-0019/2016 (H3N2), either egg-adapted (egg-derived), or wild-type sequenced.

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Table S1. Trial Design, Treatment Assignments, and Enrollment Stages

Day 0 Trial Treatment Injection

Day 28 Injectionb

Participants per Enrollment

Stage Partici-pants

Per GroupTreatment

Group Vaccine

HA Dose per Strain, μg

(H1N1/H3N2/BV/BY)

Matrix-M1 Adjuvant Dose, µg

Formulation Stage 1c

Stage 2c

Stage 3c

A

Quad-NIV

60, 60, 60, 60 50 In-clinic mix Placebo 20 20 115 155

B 60, 60, 60, 60 50 Co-form Placebo 20 60 230 310

C 60, 60, 60, 60 75 Co-form Placebo 20 20 115 155

D 60, 60, 90, 90 50 Co-form Placebo 0 20 115 135

E 60, 60, 60, 60 0 NA

2018 - 19 Licensed Seasonal Influenza Vaccine

20 60 230 310

F 2018-2019 IIV3-HD [Fluzone High-Dosea] Placebo 20 20 115 155

G 2018-2019 RIV4 [Flublok Quadrivalenta] Placebo 20 20 115 155

Total Study Participants 120 220 1035 1375Abbreviations: Bv, B Victoria lineage; BY, B Yamagata lineage; Co-form, co-formulated; HA, hemagglutinin; NA, not applicable;

IIV3-HD, trivalent high-dose inactivated influenza vaccine [Fluzone High-Dose]; RIV4, quadrivalent recombinant influenza

vaccine [Flublok Quadrivalent].

Note: All participants received 2 vaccinations by intramuscular injection in alternating deltoids on Day 0 and Day 28.aIIV3-HD and RIV4 were administered at the manufacturer’s recommended dose and volume. bOn Day 28, participants in Group E received a rescue injection with a licensed seasonal influenza vaccine (IIV3-HD; Fluzone

High-Dose); all other participants received a placebo injection to maintain trial blinding.cEnrollment was divided into 3 stages. Stage 1 enrolled a total of approximately 120 participants (approximately 20 subjects per

treatment group, excluding Group D for which no participants were enrolled in Stage 1). Stage 2 enrolled a total of

approximately 220 participants (approximately 20 [Groups A, C, D, F, and G] or 60 participants [Groups B and E] per treatment

group). The remainder of the participants (approximately 1035 subjects total, ie, 115 [Groups A, C, D, F, and G] or 230 subjects

[Groups B and E] per treatment group) were enrolled in Stage 3.

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Table S2. Schedule of Study ProceduresStudy Day: 0 3 7 28 56 90 182

Window (days): ± 1 ± 1 ± 2 ± 2 ± 7 ± 7Study informed consent X

Inclusion/exclusion criteria X

Medical/medication history X

Physical exam X Xa Xa Xa Xa

Vital signs Xb X Xb X XClinical safety laboratoryc X X

Serology X X X XPBMC for CMI Xd Xd

Study treatment injection X

Injection with a 2018-2019 licensed seasonal influenza vaccine or placeboe

X

2019-2020 licensed seasonal influenza vaccine offered to all subjectsf

Adverse event reviewf X Xg X X X X X

Concomitant medications reviewf X X X X X X X

Subject diary review Xg Xh

End of study XAbbreviation: CMI, cell-mediated immunity.

Note: Procedures shaded in light grey were performed via scripted telephone call.aIf needed, a physical examination may have been performed, based on the investigator’s discretion.bVital signs were captured pre-vaccination and between 30 and 60 minutes post-vaccination.cIncluded assessments for hematology (complete blood count [CBC] with hemoglobin, hematocrit, red blood cell [RBC] count,

platelet count, and white blood cell [WBC] count with differential) and serum chemistry (alanine aminotransferase [ALT],

aspartate aminotransferase [AST], total bilirubin, alkaline phosphatase, creatinine, and blood urea nitrogen [BUN]). dPlanned to be collected from approximately 189 subjects from 3 pre-selected sites (63 subjects per site). eOn Day 28, subjects in Group E received a rescue injection with a licensed seasonal influenza vaccine (IIV3-HD; Fluzone High-

Dose); all other subjects received a placebo injection to maintain trial blinding.fAll adverse events and concomitant medications taken were collected through Day 28; thereafter, only medically attended

events (MAEs), serious adverse events (SAEs), and significant new medical conditions (SNMCs) and medications taken for these

events were collected.

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gSubjects were asked to report any grade 3 solicited or unsolicited adverse event or SAE experienced since the last visit and may

have been asked to return to the clinic for an unscheduled visit to evaluate the event(s) at the investigator’s discretion. hThe subject diary was reviewed by the investigator and collected on Day 7 visit.

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Table S3. Summary of Baseline Demographic Data Through Day 182 – Safety Population Treatment

Group LabelA B C D E F G

HA and Matrix Content

qNIV A60/B60/M50

qNIV A60/B60/M50

qNIV A60/B60/M75

qNIV A90/B90/M50

qNIV A60/B60/M0 IIV3-HD RIV4

Formulation In-clinic mix Co-formulated NA NA NASubjects in Group (N) N = 157 N = 305 N = 156 N = 132 N = 311 N = 153 N = 151

Age (years)Mean (SD) 72.7 (5.5) 72.0 (5.5) 71.8 (5.6) 72.6 (6.1) 72.6 (6.3) 72.5 (5.5) 72.9 (5.6)Median 71.0 71.0 70.0 71.0 71.0 72.0 72.0Min, Max 65.0, 90.0 65.0, 91.0 65.0, 90.0 65.0, 91.0 65.0, 101.0 65.0, 93.0 65.0, 87.0

Gender, n (%)Male 67 (42.7) 126 (41.3) 79 (50.6) 51 (38.6) 126 (40.5) 54 (35.3) 64 (42.4)Female 90 (57.3) 179 (58.7) 77 (49.4) 81 (61.4) 185 (59.5) 99 (64.7) 87 (57.6)

Ethnicity, n (%)Hispanic or Latino

6 (3.8) 6 (2.0) 7 (4.5) 4 (3.0) 7 (2.3) 6 (3.9) 4 (2.6)

Not Hispanic or Latino

151 (96.2) 299 (98.0) 149 (95.5) 128 ( 97.0) 304 (97.7) 147 (96.1) 147 (97.4)

Race, n (%)American Indian/Alaska Native

1 (0.6) 0 (0.0) 0 (0.0) 0 (0.0) 3 (1.0) 0 (0.0) 1 (0.7)

Asian 2 (1.3) 1 (0.3) 0 (0.0) 0 (0.0) 1 (0.3) 1 (0.7) 1 (0.7)Black or African American

13 (8.3) 41 (13.4) 15 (9.6) 12 (9.1) 29 (9.3) 17 (11.1) 14 (9.3)

Native Hawaiian/Other Pacific Islander

0 (0.0) 1 (0.3) 0 (0.0) 0 (0.0) 0 (0.0) 0 (0.0) 0 (0.0)

White 138 (87.9) 261 (85.6) 140 (89.7) 120 (90.9) 277 (89.1) 133 (86.9) 135 (89.4)Other 3 (1.9) 1 (0.3) 1 (0.6) 0 (0.0) 1 (0.3) 2 (1.3) 0 (0.0)

Height (cm)Mean (SD) 167.2 (10.2) 168.6 (10.5) 168.4 (10.3) 166.9 (9.0) 166.9 (10.1) 166.7 (10.2) 167.7 (10.0)Median 166.7 167.6 168.0 165.9 165.0 165.0 166.1Min, Max 144.8, 195.0 142.2, 202.0 147.0, 199.0 148.0, 189.0 147.5, 198.1 138.4, 193.0 144.3, 191.7

Weight (kg)Mean (SD) 86.3 (20.7) 87.2 (19.3) 88.2 (19.0) 83.4 (18.6) 86.8 (19.6) 85.9 (19.8) 88.7 (22.6)Median 84.8 85.7 84.7 81.8 85.4 83.6 85.0Min, Max 46.0, 170.6 43.5, 147.3 52.4, 139.8 49.0, 132.7 47.0, 152.7 44.9, 166.9 45.4, 154.9

BMI (kg/m2)Mean (SD) 30.8 (6.8) 30.6 (6.2) 31.1 (6.3) 30.0 (6.5) 31.1 (6.3) 30.8 (6.3) 31.5 (7.5)Median 29.4 29.6 29.7 28.8 30.5 29.9 30.8Min, Max 14.2, 53.9 17.6, 52.6 19.8, 54.5 18.1, 47.3 18.4, 53.1 19.0, 50.4 18.3, 62.0

19

Abbreviations: A, influenza A strain; B, influenza B strain; BMI, body mass index; cm, centimeter; kg, kilogram; HA,

hemagglutinin; N, number of subjects who received test article at Day 0; n, number of subjects in each specified category; M,

Matrix-M1 adjuvant; m, meter; Max, maximum; Min, minimum; IIV3-HD, trivalent high-dose inactivated influenza vaccine;

qNIV, quadrivalent nanoparticle influenza vaccine; RIV4, quadrivalent recombinant influenza vaccine; SD, standard deviation.

Note: Percentages are based on the number of subjects in the safety population who received that treatment. Height and

weight are recorded at Study Day 0. Age is calculated as the lowest integer result of (Date of Study Day 0 – Date of

Birth)/365.25.

Table S4. Wild-type (wt) HAI Responses by Treatment Group, Strain, and Time Point Treatment Group

LabelA B C D E F G


qNIV A60/B60/M5

0

qNIV A60/B60/M5

0

qNIV A60/B60/M7

5

qNIV A60/B90/M5

0

qNIV A60/B60/M0 IIV3-HD RIV4

Formulation In-clinic mix Co-formulated NA NA NASubjects in Group (N) N = 149 N = 295 N = 147 N = 121 N = 290 N = 143 N = 144

Visit ParameterA/Michigan/45/2015 (H1N1) [Homologous Strain]

Day 0

GMT 50.1 47.9 55.3 48.7 48.7 50.5 44.095% CI (44.6, 56.4) (44.2, 51.9) (49.0, 62.3) (42.6, 55.8) (44.8, 52.8) (45.2, 56.4) (39.5, 49.1)

Day 28

GMT 98.9 91.3 99.1 79.5 90.3 96.9 82.195% CI (86.4, 113.1) (82.5, 101.0) (86.2, 114.0) (68.6, 92.2) (81.0, 100.5) (84.5, 111.1) (71.6, 94.2)GMFRPost/Pre 2.0 1.9 1.8 1.6 1.9 1.9 1.995% CI (1.8, 2.2) (1.8, 2.1) (1.6, 2.0) (1.5, 1.8) (1.7, 2.1) (1.7, 2.1) (1.7, 2.1)P-value <.001 <.001 <.001 <.001 <.001 <.001 <.001SCR [n, (%)] 35 [23.5] 59 [20.0] 25 [17.0] 17 [14.0] 62 [21.4] 24 [ 16.8] 31 [21.5]95% CI (16.9, 31.1) (15.6, 25.0) (11.3, 24.1) (8.4, 21.5) (16.8, 26.6) (11.1, 23.9) (15.1, 29.1)SPR [n, (%)] 143 [96.0] 272 [92.2] 142 [96.6] 111 [91.7] 267 [92.1] 133 [93.0] 129 [89.6]95% CI (91.4, 98.5) (88.5, 95.0) (92.2, 98.9) (85.3, 96.0) (88.3, 94.9) (87.5, 96.6) (83.4, 94.1)

Day 56a

GMT 83.5 72.6 82.2 61.9 90.5 78.6 65.295% CI (72.9, 95.6) (66.3, 79.7) (71.8, 94.1) (54.4, 70.3) (82.4, 99.4) (69.2, 89.4) (57.9, 73.5)GMFRPost/Pre 1.7 1.5 1.5 1.3 1.9 1.6 1.595% CI (1.5, 1.9) (1.4, 1.6) (1.3, 1.7) (1.1, 1.4) (1.7, 2.0) (1.4, 1.7) (1.3, 1.6)P-value <.001 <.001 <.001 <.001 <.001 <.001 <.001SCR [n, (%)] 18 [12.2] 27 [9.2] 11 [7.6] 4 [3.3] 42 [14.5] 7 [5.0] 11 [7.8]95% CI (7.4, 18.5) (6.1, 13.1) (3.9, 13.3) (0.9, 8.3) (10.7, 19.1) (2.0, 10.0) (4.0, 13.5)SPR [n, (%)] 123 [83.1] 225 [76.5] 121 [84.0] 92 [76.7] 246 [85.1] 119 [85.0] 107 [75.9]95% CI (76.1, 88.8) (71.3, 81.3) (77.0, 89.6) (68.1, 83.9) (80.5, 89.0) (78.0, 90.5) (68.0, 82.7)

Day 182a

GMT 66.3 58.4 65.6 53.6 71.6 65.6 53.995% CI (58.0, 75.9) (53.4, 63.9) (57.2, 75.3) (47.0, 61.2) (65.4, 78.4) (58.8, 73.1) (47.8, 60.7)GMFRPost/Pre 1.3 1.2 1.2 1.1 1.5 1.3 1.295% CI (1.2, 1.5) (1.1, 1.3) (1.1, 1.3) (1.0, 1.2) (1.4, 1.6) (1.2, 1.4) (1.1, 1.4)P-value <.001 <.001 0.001 0.127 <.001 <.001 <.001SCR [n, (%)] 5 [3.5] 11 [3.8] 4 [2.9] 0 [ 0] 22 [7.8] 3 [2.1] 3 [2.1]95% CI (1.1, 7.9) (1.9, 6.7) (0.8, 7.2) (0.0, 3.1) (5.0, 11.6) (0.4, 6.1) (0.4, 6.1)

20

SPR [n, (%)] 102 [70.8] 176 [61.1] 97 [69.3] 67 [57.3] 198 [70.5] 97 [68.8] 84 [59.6]95% CI (62.7, 78.1) (55.2, 66.8) (60.9, 76.8) (47.8, 66.4) (64.8, 75.7) (60.5, 76.3) (51.0, 67.7)

A/Singapore/INFIMH-16-0019/2016 (H3N2) [Homologous Strain]

Day 0

GMT 21.4 22.0 22.8 23.4 21.3 23.2 19.595% CI (18.9, 24.3) (20.0, 24.1) (19.7, 26.3) (20.2, 27.1) (19.3, 23.5) (20.4, 26.4) (17.1, 22.2)

Day 28


Day 56a

GMT 51.2 50.1 52.6 48.1 53.4 38.9 49.395% CI (44.1, 59.4) (45.4, 55.3) (45.3, 61.0) (41.2, 56.1) (48.1, 59.2) (33.7, 45.1) (42.0, 57.9)GMFRPost/Pre 2.4 2.3 2.3 2.1 2.5 1.7 2.595% CI (2.1, 2.7) (2.1, 2.5) (2.0, 2.6) (1.8, 2.4) (2.3, 2.8) (1.5, 1.9) (2.1, 3.0)P-value <.001 <.001 <.001 <.001 <.001 <.001 <.001SCR [n, (%)] 34 [23.0] 59 [20.1] 29 [20.1] 19 [15.8] 63 [21.8] 13 [ 9.3] 37 [26.2]95% CI (16.5, 30.6) (15.6, 25.1) (13.9, 27.6) (9.8, 23.6) (17.2, 27.0) (5.0, 15.4) (19.2, 34.3)SPR [n, (%)] 88 [59.5] 172 [58.5] 88 [61.1] 74 [61.7] 176 [60.9] 63 [45.0] 81 [57.4]95% CI (51.1, 67.4) (52.6, 64.2) (52.6, 69.1) (52.4, 70.4) (55.0, 66.6) (36.6, 53.6) (48.8, 65.7)

Day 182a


B/Colorado/06/2017 (Victoria Lineage) [Homologous Strain]

Day 0GMT 47.6 46.4 48.5 52.7 47.5 52.0 42.995% CI (43.1, 52.6) (42.8, 50.2) (44.0, 53.6) (46.7, 59.4) (43.9, 51.3) (46.1, 58.5) (38.5, 47.7)

Day 28


Day 56a

GMT 67.8 64.3 67.9 72.3 74.9 70.9 60.795% CI (60.2, 76.4) (58.6, 70.6) (60.2, 76.5) (62.8, 83.3) (69.7, 80.4) (62.2, 80.7) (53.6, 68.7)GMFRPost/Pre 1.4 1.4 1.4 1.4 1.6 1.4 1.4

21

95% CI (1.3, 1.6) (1.3, 1.5) (1.2, 1.5) (1.2, 1.5) (1.5, 1.7) (1.2, 1.5) (1.3, 1.6)P-value <.001 <.001 <.001 <.001 <.001 <.001 <.001SCR [n, (%)] 7 [4.7] 17 [5.8] 4 [2.8] 6 [5.0] 14 [4.8] 8 [5.7] 6 [4.3]95% CI (1.9, 9.5) (3.4, 9.1) (0.8, 7.0) (1.9, 10.6) (2.7, 8.0) (2.5, 10.9) (1.6, 9.0)SPR [n, (%)] 108 [73.0] 188 [63.9] 104 [72.2] 89 [74.2] 233 [80.6] 103 [73.6] 92 [65.2]95% CI (65.1, 79.9) (58.2, 69.4) (64.2, 79.4) (65.4, 81.7) (75.6, 85.0) (65.5, 80.7) (56.8, 73.1)

Day 182a


B/Phuket/3073/2013 (Yamagata Lineage) [Homologous Strain]

Day 0

GMT 50.4 47.7 47.6 53.6 48.8 52.9 46.995% CI (44.7, 56.7) (44.4, 51.3) (42.9, 52.9) (46.9, 61.2) (45.2, 52.7) (47.3, 59.1) (42.1, 52.2)

Day 28

GMT 108.5 101.7 104.9 113.8 87.5 64.5 102.095% CI (96.2, 122.4) (93.3, 110.8) (92.4, 119.2) (97.6, 132.6) (79.5, 96.4) (57.3, 72.6) (88.6,

117.4)GMFRPost/Pre 2.2 2.1 2.2 2.1 1.8 1.2 2.295% CI (1.9, 2.4) (2.0, 2.3) (2.0, 2.5) (1.8, 2.5) (1.6, 2.0) (1.1, 1.3) (1.9, 2.5)P-value <.001 <.001 <.001 <.001 <.001 <.001 <.001SCR [n, (%)] 40 [26.8] 76 [25.8] 41 [27.9] 41 [33.9] 57 [19.7] 5 [3.5] 40 [27.8]95% CI (19.9, 34.7) (20.9, 31.2) (20.8, 35.9) (25.5, 43.0) (15.2, 24.7) (1.1, 8.0) (20.6, 35.8)SPR [n, (%)] 147 [98.7] 290 [98.3] 144 [98.0] 117 [96.7] 271 [93.4] 126 [88.1] 136 [94.4]95% CI (95.2, 99.8) (96.1, 99.4) (94.2, 99.6) (91.8, 99.1) (90.0, 96.0) (81.6, 92.9) (89.3, 97.6)

Day 56a

GMT 96.8 92.3 96.6 102.0 87.0 60.9 85.595% CI (85.5, 109.5) (84.1, 101.2) (84.4, 110.6) (87.1, 119.4) (79.1, 95.7) (53.6, 69.2) (73.7, 99.1)GMFRPost/Pre 1.9 1.9 2.0 1.9 1.8 1.2 1.895% CI (1.7, 2.2) (1.8, 2.1) (1.8, 2.3) (1.7, 2.2) (1.6, 1.9) (1.1, 1.3) (1.6, 2.1)P-value <.001 <.001 <.001 <.001 <.001 <.001 <.001SCR [n, (%)] 31 [20.9] 48 [16.3] 31 [21.5] 30 [25.0] 44 [15.2] 3 [2.1] 26 [18.4]95% CI (14.7, 28.4) (12.3, 21.1) (15.1, 29.1) (17.5, 33.7) (11.3, 19.9) (0.4, 6.1) (12.4, 25.8)SPR [n, (%)] 140 [ 94.6] 273[ 92.9] 135 [ 93.8] 109 [ 90.8] 262 [ 90.7] 114 [ 81.4] 124 [ 87.9]95% CI (89.6, 97.6) (89.3, 95.5) (88.5, 97.1) (84.2, 95.3) (86.7, 93.8) (74.0, 87.5) (81.4, 92.8)

Day 182a

GMT 66.0 63.0 67.7 69.4 63.5 57.3 63.095% CI (59.9, 72.9) (58.7, 67.7) (61.1, 75.0) (61.7, 78.1) (59.0, 68.4) (52.2, 63.1) (56.7, 69.9)GMFRPost/Pre 1.3 1.3 1.4 1.3 1.3 1.1 1.495% CI (1.2, 1.4) (1.2, 1.4) (1.3, 1.6) (1.2, 1.5) (1.2, 1.4) (1.0, 1.2) (1.2, 1.5)P-value <.001 <.001 <.001 <.001 <.001 0.014 <.001SCR [n, (%)] 8 [5.6] 8 [2.8] 6 [4.3] 5 [4.3] 9 [3.2] 0 [0] 4 2.8]95% CI (2.4, 10.7) (1.2, 5.4) (1.6, 9.1) (1.4, 9.7) (1.5, 6.0) (0.0, 2.6) (0.8, 7.1)SPR [n, (%)] 117 [81.3] 219 [76.0] 109 [77.9] 95 [81.2] 210 [74.7] 104 [73.8] 106 [75.2]95% CI (73.9, 87.3) (70.7, 80.9) (70.1, 84.4) (72.9, 87.8) (69.2, 79.7) (65.7, 80.8) (67.2, 82.1)

A/Switzerland/9715293/2013 (H3N2) [Drift Strain]Day GMT 54.6 60.0 60.9 60.6 58.6 62.3 54.8

22

0 95% CI (47.8, 62.4) (54.3, 66.2) (52.7, 70.3) (52.1, 70.5) (52.8, 65.1) (54.3, 71.6) (47.8, 62.9)

Day 28

GMT 146.8 160.4 154.8 137.9 122.4 133.4 158.895% CI (124.0,

173.9)(143.9, 178.7)

(132.7, 180.7)

(117.2, 162.2)

(109.1, 137.3)

(111.2, 160.0)

(132.2, 190.9)

GMFRPost/Pre 2.7 2.7 2.5 2.3 2.1 2.1 2.995% CI (2.3, 3.1) (2.4, 3.0) (2.2, 2.9) (2.0, 2.7) (1.9, 2.3) (1.9, 2.4) (2.5, 3.4)P-value <.001 <.001 <.001 <.001 <.001 <.001 <.001SCR [n, (%)] 54 [36.2] 105 [35.6] 43 [29.3] 35 [28.9] 72 [24.8] 31 [21.7] 57 [39.6]95% CI (28.5, 44.5) (30.1, 41.3) (22.0, 37.3) (21.0, 37.9) (20.0, 30.2) (15.2, 29.3) (31.5, 48.1)SPR [n, (%)] 142 [95.3] 289 [98.0] 143 [97.3] 118 [97.5] 269 [92.8] 132 [92.3] 134 [93.1]95% CI (90.6, 98.1) (95.6, 99.3) (93.2, 99.3) (92.9, 99.5) (89.1, 95.5) (86.7, 96.1) (87.6, 96.6)

Day 56a

GMT 120.1 127.1 124.6 113.9 132.4 108.3 123.895% CI (103.0,

140.1)(115.3, 140.2)

(107.9, 143.8)

(97.1, 133.5) (118.8, 147.7)

(92.4, 126.9) (104.8, 146.2)

GMFRPost/Pre 2.2 2.1 2.0 1.9 2.3 1.7 2.395% CI (1.9, 2.5) (1.9, 2.3) (1.8, 2.3) (1.6, 2.2) (2.1, 2.5) (1.5, 1.9) (1.9, 2.6)P-value <.001 <.001 <.001 <.001 <.001 <.001 <.001SCR [n, (%)] 38 [25.7] 74 [25.2] 28 [19.4] 24 [20.0] 79 [27.3] 17 [12.1] 37 [26.2]95% CI (18.9, 33.5) (20.3, 30.5) (13.3, 26.9) (13.3, 28.3) (22.3, 32.9) (7.2, 18.7) (19.2, 34.3)SPR [n, (%)] 141 [95.3] 288 [98.0] 136 [94.4] 113 [94.2] 273 [94.5] 132 [94.3] 129 [91.5]95% CI (90.5, 98.1) (95.6, 99.2) (89.3, 97.6) (88.4, 97.6) (91.2, 96.8) (89.1, 97.5) (85.6, 95.5)

Day 182a

GMT 89.9 92.0 95.4 82.0 97.9 81.2 93.895% CI (76.7, 105.5) (82.6, 102.4) (81.2, 112.1) (69.7, 96.4) (87.2, 110.0) (69.5, 94.8) (79.3,

111.0)GMFRPost/Pre 1.6 1.5 1.5 1.4 1.7 1.3 1.795% CI (1.4, 1.9) (1.4, 1.7) (1.4, 1.7) (1.2, 1.6) (1.5, 1.8) (1.2, 1.4) (1.5, 1.9)P-value <.001 <.001 <.001 <.001 <.001 <.001 <.001SCR [n, (%)] 18 [12.5] 23 [8.0] 9 [6.4] 8 [6.8] 28 [10.0] 6 [ 4.3] 18 [12.8]95% CI (7.6, 19.0) (5.1, 11.7) (3.0, 11.9) (3.0, 13.0) (6.7, 14.1) (1.6, 9.0) (7.7, 19.4)SPR [n, (%)] 115 [79.9] 233 [80.9] 112 [80.0] 93 [79.5] 226 [80.4] 103 [73.0] 109 [77.3]95% CI (72.4, 86.1) (75.9, 85.3) (72.4, 86.3) (71.0, 86.4) (75.3, 84.9) (64.9, 80.2) (69.5, 83.9)

A/Wisconsin/19/2017 (H3N2) [Drift Strain]

Day 0

GMT 21.7 23.1 24.0 23.9 22.1 24.5 20.795% CI (19.2, 24.6) (21.1, 25.3) (20.7, 27.7) (20.9, 27.3) (20.1, 24.4) (21.6, 27.7) (18.1, 23.6)

Day 28

GMT 61.1 63.2 63.0 58.2 50.1 46.1 64.395% CI (51.8, 72.0) (56.3, 70.9) (53.5, 74.2) (48.9, 69.2) (44.4, 56.5) (38.7, 55.1) (53.5, 77.2)GMFRPost/Pre 2.8 2.7 2.6 2.4 2.3 1.9 3.195% CI (2.4, 3.3) (2.5, 3.0) (2.3, 3.0) (2.1, 2.9) (2.0, 2.5) (1.7, 2.1) (2.6, 3.7)P-value <.001 <.001 <.001 <.001 <.001 <.001 <.001SCR [n, (%)] 62 [41.6] 104 [35.3] 52 [35.4] 37 [30.6] 87 [30.0] 26 [18.2] 62 [43.1]95% CI (33.6, 50.0) (29.8, 41.0) (27.7, 43.7) (22.5, 39.6) (24.8, 35.6) (12.2, 25.5) (34.8, 51.6)SPR [n, (%)] 113 [75.8] 23 1[78.3] 116 [78.9] 91 [75.2] 199 [68.6] 93 [65.0] 105 [72.9]95% CI (68.2, 82.5) (73.2, 82.9) (71.4, 85.2) (66.5, 82.6) (62.9, 73.9) (56.6, 72.8) (64.9, 80.0)

Day 56a

GMT 56.0 57.4 60.6 53.4 60.6 41.7 55.795% CI (47.6, 65.9) (51.6, 63.9) (51.4, 71.5) (45.2, 63.0) (54.2, 67.7) (35.4, 49.2) (46.6, 66.6)GMFRPost/Pre 2.6 2.5 2.5 2.2 2.7 1.7 2.795% CI (2.2, 3.0) (2.3, 2.7) (2.2, 2.8) (1.9, 2.6) (2.5, 3.0) (1.5, 1.9) (2.3, 3.2)P-value <.001 <.001 <.001 <.001 <.001 <.001 <.001

23

SCR [n, (%)] 28 [18.9] 49 [16.7] 24 [16.7] 15 [12.5] 59 [20.4] 8 [5.7] 31 [22.0]95% CI (13.0, 26.2) (12.6, 21.4) (11.0, 23.8) (7.2, 19.8) (15.9, 25.5) (2.5, 10.9) (15.5, 29.7)SPR [n, (%)] 79 [53.4] 156 [53.1] 83 [57.6] 66 [55.0] 166 [57.4] 59 [42.1] 76 [53.9]95% CI (45.0, 61.6) (47.2, 58.9) (49.1, 65.8) (45.7, 64.1) (51.5, 63.2) (33.9, 50.8) (45.3, 62.3)

Day 182a


Abbreviations: CI, confidence interval; GMT, geometric mean titer; GMRPost/Pre, ratio of GMTs at point Day 28/Day 0; N,

number of subjects in per protocol (PP) population; n, number of subjects with non-missing hemagglutination-inhibition

(HAI) titer results at each visit; wt, wild-type.

aNote: Since Day 56/182 samples were tested separately from Day 0 and Day 28 samples, Day 56/182 titers were adjusted

for the long-term assay variability. The adjustment was based on re-testing of randomly selected subset 50 subjects of Day 0

samples concurrently with Day 56/182 samples.

GMT was defined as the antilog of the mean of the log-transformed titer values for a given treatment group and time point.

Individual antibody values recorded as below the lower limit of quantitation (LLOQ) were set to half LLOQ.

GMFRPost/Pre was defined as the ratio of 2 geometric mean titers within treatment group at 2 different time points between

post-vaccination (Day 28) and pre-vaccination (Day 0). The 95% CI and P-value were obtained by paired t-test of GMR = 1.

Individual antibody values recorded as below the LLOQ were set to half LLOQ.

SCR, seroconversion rate, was defined as percentage of subjects with either a baseline HAI titer <1:10 and a post-vaccination

titer ≥1:40, or a baseline HAI titer ≥1:10 and a 4-fold increase in post-vaccination HAI titer relative to baseline. Percentages

are based on the number of subjects with non-missing HAI titer values in the PP population who received that treatment.

Clopper-Pearson method was applied to calculate the proportion CI.

SPR, seroprotection rate, was defined as percentage of subjects with an HAI titer ≥1:40. Clopper-Pearson method was

applied to calculate the proportion CI.. Percentages were based on the number of subjects with non-missing measurements

in the PP population within treatment group.

24

25

Table S5A. Geometric Mean Counts and Geometric Mean Fold Rise of Specific CD4 T-Cells Expressing Double and Triple Cytokines per 106 CD4 T-Cells – Intent-to-Treat Population

Treatment Group Label A B C D E F GHA and Matrix Content qNIV

A60/B60/M50qNIV

A60/B60/M50qNIV

A60/B60/M75qNIV

A60/B90/M50qNIV

A60/B60/M0 IIV3-HD RIV4

Formulation Bed side Co-formulated NA NA NASubjects in Group (N) N = 157 N = 305 N = 156 N = 132 N = 311 N = 153 N = 151

Visit Parameter

A/Michigan/45/2015 (H1N1) [Homologous Strain]Subjects Tested (n) 0 4 4 0 0 3 7

Day 0

DB

Cyto

kine

GMC NC 51.6 348.4 NC NC 8.3 10.795% CI (NC, NC) (0.7, 3669.7) (54.5, 2229.4) (NC, NC) (NC, NC) (0.0, 76647.8) (0.7, 172.9)

TP

Cyto

kine

GMC NC 14.9 1623.1 NC NC 5.5 9.395% CI (NC, NC) (0.1, 2260.5) (194.7,

13531.6)(NC, NC) (NC, NC) (0.0, 8758.5) (0.7, 122.2)

Subjects Tested (n) 0 4 4 0 0 3 7

Day 7

DB C

ytok

ine

GMC NC 65.5 4190.7 NC NC 55.6 163.2

95% CI (NC, NC) (0.3, 10169.6) (585.5, 29994.4)

(NC, NC) (NC, NC) (0.0, 625150.1) (17.5, 1525.0)

GMFRPost/

Pre

NC 1.3 12.0 NC NC 6.7 15.2

95% CI (NC, NC) (0.2, 9.3) (2.1, 69.3) (NC, NC) (NC, NC) (0.0, 2684.8) (0.3, 680.7)NC 0.728 0.020 NC NC 0.307 0.130

TP C

ytok

ine

GMC NC 14.9 1623.1 NC NC 5.4 122.6

95% CI (NC, NC) (0.1, 2260.5) (194.7, 13531.6)

(NC, NC) (NC, NC) (0.0, 7868.5) (53.3, 281.8)

GMFRPost/

Pre

NC 4.8 23.1 NC NC 1.0 13.2

95% CI (NC, NC) (0.0, 536.7) (0.5, 976.1) (NC, NC) (NC, NC) (0.9, 1.1) (0.7, 240.5)p-value NC 0.368 0.076 NC NC 0.423 0.072

A/Singapore/INFIMH-16-0019/2016 (H3N2) [Homologous Strain]Subjects Tested (n) 16 15 11 16 17 11 12

Day 0

DB C

ytok

ine GMC 24.2 87.8 15.8 38.7 17.0 10.2 8.8

95% CI (6.9, 84.5) (17.9, 431.5) (1.8, 142.0) (8.9, 167.1) (4.6, 63.5) (1.6, 67.6) (1.6, 49.8)

TP

Cyto

kine

GMC 2.7 8.6 5.3 6.9 2.8 3.9 6.795% CI (1.0, 7.4) (1.9, 39.1) (0.8, 36.1) (1.7, 27.9) (1.2, 6.8) (0.8, 20.4) (1.4, 31.9)

Subjects Tested (n) 15 15 11 16 18 12 13Day 7

DB C

ytok

ine

GMC 147.1 548.8 837.6 876.3 77.7 27.4 100.3

95% CI (31.6, 684.7) (144.0, 2091.6)

(174.0, 4031.6)

(385.6, 1991.4)

(25.7, 234.7) (2.9, 257.8) (16.4, 615.1)

GMFRPost/

Pre

6.5 6.3 52.9 22.7 4.5 2.3 9.0

95% CI (1.1, 39.3) (0.5, 74.8) (5.0, 553.8) (7.9, 64.7) (1.3, 15.8) (0.2, 26.6) (0.7, 114.5)p-value 0.042 0.136 0.004 <.001 0.023 0.463 0.084

TP

Cyto

ki GMC 22.4 275.8 265.5 166.0 10.1 7.6 34.795% CI (3.9, 130.3) (69.7, 1091.0) (39.6, 1780.2) (42.5, 648.9) (2.9, 34.4) (1.1, 52.6) (5.3, 225.7)

26

ne

GMFRPost/

Pre

7.9 32.2 50.2 24.1 4.1 2.3 4.0

95% CI (0.8, 76.1) (3.1, 328.5) (5.1, 490.8) (7.4, 78.8) (1.1, 14.9) (0.3, 16.4) (0.7, 22.1)p-value 0.070 0.006 0.003 <.001 0.036 0.354 0.098

B/Colorado/06/2017 (Victoria Lineage) [Homologous Strain]Subjects Tested (n) 16 16 14 15 17 11 12

Day 0

DB

Cyto

kine

GMC 172.1 91.3 45.6 118.0 69.5 53.0 111.995% CI (102.4, 289.1) (20.2, 413.4) (9.2, 226.4) (31.4, 443.4) (20.9, 231.6) (8.0, 350.1) (25.0, 501.6)

TP

Cyto

kine

GMC 20.4 13.1 9.1 17.8 17.2 16.3 35.995% CI (6.2, 67.3) (3.0, 56.4) (1.9, 44.6) (4.1, 77.0) (5.6, 53.0) (2.5, 105.2) (6.2, 207.6)


Day 7

DB C

ytok

ine

GMC 804.5 1065.5 2924.9 1386.7 272.8 433.1 602.9

95% CI (454.5, 1423.9)

(318.7, 3561.7)

(1477.4, 5790.4)

(616.6, 3118.8)

(121.2, 614.2) (113.3, 1656.1) (165.6, 2194.7)

GMFRPost/

Pre

4.7 11.7 64.1 11.8 3.8 7.5 4.7

95% CI (2.5, 8.7) (1.9, 70.2) (10.5, 391.4) (3.0, 46.5) (0.9, 15.3) (1.3, 41.9) (1.5, 15.4)p-value <.001 0.011 <.001 0.002 0.059 0.026 0.014

TP C

ytok

ine

GMC 305.9 463.6 1419.6 419.3 41.7 146.8 166.9

95% CI (159.6, 586.3) (145.5, 1477.2)

(685.5, 2940.0)

(153.7, 1143.3)

(13.8, 125.4) (31.4, 687.2) (36.5, 763.2)

GMFRPost/

Pre

14.5 35.4 155.8 23.6 3.0 7.7 4.0

95% CI (3.9, 54.0) (5.7, 218.8) (26.8, 904.1) (4.1, 137.2) (0.7, 13.8) (1.1, 55.7) (0.4, 45.8)p-value <.001 <.001 <.001 0.002 0.144 0.045 0.236

A/Wisconsin/19/2017 (H3N2) [Drift strain]Subjects Tested (n) 13 11 8 12 14 7 9

Day 0

DB C

ytok

ine GMC 35.2 110.9 18.8 19.4 51.7 34.9 31.3

95% CI (9.0, 137.3) (18.8, 655.0) (1.3, 271.5) (3.9, 95.9) (14.2, 188.6) (1.6, 781.4) (4.3, 229.5)

TP

Cyto

kine

GMC 2.1 6.6 2.0 1.5 6.9 5.4 23.095% CI (0.7, 6.5) (1.0, 42.1) (0.4, 10.2) (0.6, 4.0) (1.7, 27.5) (0.4, 79.4) (3.7, 143.2)


Day 7 DB C

ytok

ine

GMC 89.3 1098.8 1834.3 426.9 106.3 160.9 165.5

95% CI (10.7, 741.5) (456.4, 2645.5)

(388.9, 8651.3)

(93.0, 1959.2) (33.1, 341.4) (25.2, 1028.5) (21.6, 1268.1)

GMFRPost/

Pre

2.5 13.6 97.4 36.5 2.0 4.7 4.1

95% CI (0.3, 18.1) (2.2, 82.7) (2.1, 4541.5) (4.6, 288.7) (0.3, 13.7) (0.4, 49.5) (1.3, 12.8)p-value 0.335 0.010 0.026 0.003 0.432 0.162 0.020

TP C

ytok

ine

GMC 11.1 249.1 122.3 129.0 34.3 13.7 74.895% CI (1.6, 77.3) (45.2, 1373.6) (4.1, 3652.8) (22.1, 751.2) (10.8, 108.7) (1.2, 152.7) (12.7, 440.6)GMFRPost/

Pre

5.0 88.4 61.3 125.4 4.6 1.8 2.5

95% CI (0.6, 38.4) (14.8, 530.3) (1.8, 2132.2) (21.9, 719.3) (0.7, 30.6) (0.0, 201.8) (0.7, 8.8)p-value 0.112 <.001 0.029 <.001 0.103 0.770 0.128

27

Abbreviations: CI, confidence interval; GMC, geometric mean count; GMFRPost/Pre, geometric mean count fold rise; HA,

hemagglutinin; IIV3-HD, trivalent high-dose inactivated influenza vaccine; qNIV, quadrivalent recombinant nanoparticle

influenza vaccine; RIV4, quadrivalent recombinant influenza vaccine;

Cell-mediated immune (CMI) responses were measured by intracellular cytokine staining (ICCS). Counts of peripheral blood

CD4+ T-cells producing interleukin-2 (IL-2), interferon gamma (IFN-γ), and/or tumor necrosis factor alpha (TNF-α) cytokines

were measured following in vitro re-stimulation with vaccine-homologous (A/Singapore/FIMH-16-0019/2016 [H3N2];

A/Michigan/45/2015 [H1N1]; B/Colorado/06/2017 [Victoria]), or drifted (A/Wisconsin/19/2017 [H3N2]) strain-specific

recombinant wild-type sequence HAs.

GMCs were calculated for double or triple cytokine producing (ie, double cytokine was any 2 of: IFN-γ, TNF-α, or IL-2; triple

cytokine was all of: IFN-γ, TNF-α, and IL-2) strain-specific CD4+ T cells individually by each strain for all samples tested across

the 4 strains evaluated, using PBMCs obtained from a subgroup of subjects on Day 0 (pre-vaccination) and Day 7. The lower

limit of quantitation (LLOQ) was defined as 1 if the count was 0. GMFRPost/Pre, was calculated as the Day 7/Day 0 ratio of GMCs

of double or triple cytokine producing strain-specific CD4+ T-cells for each treatment group.

Note that the number of strains tested for a given participant’s sample was dependent on the number of cells available;

thus, not all samples could be tested across all 4 strains.

28

Table S5B. Baseline Adjusted Ratio of Geometric Mean Counts of Specific CD4 T-Cells Expressing Double and Triple Cytokines per 106 CD4 T-Cells in Quad-NIV Groups B and C vs IIV3-HD – Intent-to-Treat Population

Treatment Group Labels B vs F C vs FHA and Matrix Content qNIV A60/B60/M50

Co-formulatedqNIV A60/B60/M75

Co-formulatedComparator IIV3-HD IIV3-HD

ParameterA/Michigan/45/2015 (H1N1) [Homologous Strain]

Counts of Double Cytokine

n1, n2 4, 3 4, 3GMCR 0.23 4.0895% CI (0.01, 7.08) (0.07, 228.02)P-value 0.414 0.498

Counts of Triple Cytokine

n1, n2 4, 3 4, 3GMCR 4.39 66.6495% CI (0.07, 271.58) (2.85, 1558.50)P-value 0.487 0.047



n1, n2 15, 11 11, 11GMCR 23.62 30.7595% CI (2.53, 220.48) (3.38, 279.45)P-value 0.024 0.015


n1, n2 15, 11 11, 11GMCR 28.75 25.4695% CI (4.14, 199.92) (3.16, 205.25)P-value 0.007 0.015



n1, n2 16, 11 14, 11GMCR 2.37 7.5295% CI (0.54, 10.38) (2.34, 24.13)P-value 0.326 0.007


n1, n2 16, 11 14, 11GMCR 3.83 12.3095% CI (0.81, 18.00) (3.30, 45.91)P-value 0.151 0.003



n1, n2 10, 7 8, 7GMCR 5.77 11.5695% CI (1.52, 21.87) (1.50, 89.26)P-value 0.036 0.054


n1, n2 10, 7 8, 7GMCR 37.45 9.8695% CI (3.83, 366.07) (0.30, 319.00)P-value 0.014 0.263

29

Abbreviations: CI, confidence interval; GMC, geometric mean count; GMCR, ratio of GMCs between treatment arms at Day 7;

HA, hemagglutinin; IIV3-HD, trivalent high-dose inactivated influenza vaccine; qNIV, quadrivalent recombinant nanoparticle

influenza vaccine; RIV4, quadrivalent recombinant influenza vaccine.







cytokine was all of: IFN-γ, TNF-α, and IL-2) strain-specific CD4+ T-cells individually by each strain for all samples tested across


limit of quantitation (LLOQ) was defined as 1 if the count was 0. GMFRPost/Pre, was calculated as the within treatment group

Day 7/Day 0 ratio of GMCs of double or triple cytokine producing strain-specific CD4+ T-cells for each treatment group.

GMCR was defined as the ratio of GMCs for a comparison between 2 specified treatment groups at Day 7 of double or triple

cytokine producing strain-specific CD4+ T-cells.



Note: For GMCR, a mixed effects model with treatment group and baseline counts as covariate was performed. The ratios of

geometric least square (LS) means and 90% CIs for the ratios are calculated by back transforming the mean differences and

90% confidence limits for the differences of log10 transformed total counts of triple cytokine or double cytokine between 2

specified treatment groups.

Note: Lower limit of quantitation (LLOQ) is set as 1; if the cytokine count was 0, log10 scale of cytokine counts is recorded as

log10 scale LLOQ.

30

Table S5C. Baseline Adjusted Ratio of Geometric Mean Counts of Specific CD4 T-Cells Expressing Double and Triple Cytokines per 106 CD4 T-Cells in Quad-NIV Groups B and C vs RIV4 – Intent-to-Treat Population

Treatment Group Labels B vs G C vs GHA and Matrix Content qNIV A60/B60/M50

Co-formulatedqNIV A60/B60/M75

Co-formulatedComparator RIV4 RIV4

ParameterA/Michigan/45/2015 (H1N1) [Homologous Strain]


n1, n2 4, 7 4, 7GMCR 0.29 31.5295% CI (0.01, 7.61) (1.29, 772.36)P-value 0.502 0.080


n1, n2 4, 7 4, 7GMCR 0.13 14.1295% CI (0.01, 1.59) (3.42, 58.33)P-value 0.169 0.008



n1, n2 15, 12 11, 12GMCR 11.05 9.9295% CI (1.60, 76.10) (1.36, 72.60)P-value 0.044 0.061


n1, n2 15, 12 11, 12GMCR 9.89 10.9695% CI (1.53, 63.91) (1.50, 80.31)P-value 0.046 0.051



n1, n2 16, 12 14, 12GMCR 2.13 6.6395% CI (0.53, 8.61) (2.12, 20.72)P-value 0.366 0.009


n1, n2 16, 12 14, 12GMCR 3.25 9.6295% CI (0.66, 15.91) (2.37, 39.02)P-value 0.217 0.011



n1, n2 10, 9 8, 9GMCR 5.60 15.7395% CI (1.39, 22.56) (1.86, 132.91)P-value 0.046 0.039


n1, n2 10, 9 8, 9GMCR 17.30 9.6695% CI (3.28, 91.21) (0.43, 217.05)P-value 0.009 0.220

Abbreviations: CI, confidence interval; GMC, geometric mean count; GMCR, ratio of GMCs between treatment arms at Day 7;

31

HA, hemagglutinin; IIV3-HD, trivalent high-dose inactivated influenza vaccine; qNIV, quadrivalent recombinant nanoparticle

influenza vaccine; RIV4, quadrivalent recombinant influenza vaccine.;







cytokine was all of: IFN-γ, TNF-α, and IL-2) strain-specific CD4+ T-cells individually by each strain for all samples tested across


limit of quantitation (LLOQ) was defined as 1 if the count was 0. GMFRPost/Pre, was calculated as the within treatment group

Day 7/Day 0 ratio of GMCs of double or triple cytokine producing strain-specific CD4+ T-cells for each treatment group.

GMCR was defined as the ratio of GMCs for a comparison between 2 specified treatment groups at Day 7 of double or triple

cytokine producing strain-specific CD4+ T-cells.



Note: For GMCR, a mixed effects model with treatment group and baseline counts as covariate was performed. The ratios of

geometric least square (LS) means and 90% CIs for the ratios are calculated by back transforming the mean differences and

90% confidence limits for the differences of log10 transformed total counts of triple cytokine or double cytokine between 2

specified treatment groups.

Note: Lower limit of quantitation (LLOQ) is set as 1; if the cytokine count was 0, log10 scale of cytokines counts is recorded as

log10 scale LLOQ.

32

Table S6. Summary of All and Severe Solicited Adverse Events From Day 0 Post-vaccination to Day 6, Inclusive – Safety Population

Treatment Group Label

A B C D E F G


qNIV A60/B60/M50

qNIV A60/B60/M50

qNIV A60/B60/M75

qNIV A60/B90/M50

qNIV A60/B60/M0

IIV3-HD RIV4

Formulation Bedside Co-formulated NA NA NASubjects in Group

(N)N =157 N = 305 N = 156 N = 132 N = 311 N = 153 N = 151

n (% of subjects)95% CI

All solicited AEs 61 (38.9) 99 (32.5) 59 (37.8) 39 (29.5) 85 (27.3) 58 (37.9) 56 (37.1)All severe solicited AEs 3 (1.9) 10 (3.3) 5 (3.2) 2 (1.5) 4 (1.3) 2 (1.3) 4 (2.6)

Solicited Local AEsAll local AEs 30 (19.1) 74 (24.3) 34 (21.8) 22 (16.7) 40 (12.9) 40 (26.1) 22 (14.6)All severe local AEs 1 (0.6) 2 (0.7) 0 (0.0) 0 (0.0) 1 (0.3) 0 (0.0) 0 (0.0)

Bruising 2 (1.3) 11 (3.6) 5 (3.2) 3 (2.3) 6 (1.9) 4 (2.6) 5 (3.3)Pain 22 (14.0) 59 (19.3) 30 (19.2) 15 (11.4) 32 (10.3) 32 (20.9) 14 (9.3)Redness 12 (7.6) 21 (6.9) 8 (5.1) 6 (4.5) 11 (3.5) 10 (6.5) 4 (2.6)

Severe 1 (0.6) 0 (0.0) 0 (0.0) 0 (0.0) 1 (0.3) 0 (0.0) 0 (0.0)Swelling 6 (3.8) 31 (10.2) 11 (7.1) 11 (8.3) 16 (5.1) 12 (7.8) 7 (4.6)

Severe 0 (0.0) 2 (0.7) 0 (0.0) 0 (0.0) 1 (0.3) 0 (0.0) 0 (0.0)General Systemic AEs

All systemic AEs 42 (26.8) 63 (20.7) 45 (28.8) 27 (20.5) 65 (20.9) 37 (24.2) 39 (25.8)All severe systemic AEs

2 (1.3) 9 (3.0) 5 (3.2) 2 (1.5) 3 (1.0) 2 (1.3) 4 (2.6)

Chills 6 (3.8) 10 (3.3) 7 (4.5) 3 (2.3) 8 (2.6) 5 (3.3) 2 (1.3)Severe 1 (0.6) 0 (0.0) 1 (0.6) 0 (0.0) 0 (0.0) 1 (0.7) 0 (0.0)

Fatigue 17 (10.8) 25 (8.2) 13 (8.3) 10 (7.6) 16 (5.1) 16 (10.5) 10 (6.6)Severe 0 (0.0) 2 (0.7) 2 (1.3) 0 (0.0) 2 (0.6) 1 (0.7) 1 (0.7)

Headache 16 (10.2) 23 (7.5) 23 (14.7) 13 (9.8) 23 (7.4) 14 (9.2) 14 (9.3)Severe 0 (0.0) 2 (0.7) 2 (1.3) 1 (0.8) 0 (0.0) 2 (1.3) 3 (2.0)

Joint pain 8 (5.1) 16 (5.2) 12 (7.7) 5 (3.8) 12 (3.9) 12 (7.8) 10 (6.6)Severe 0 (0.0) 0 (0.0) 1 (0.6) 0 (0.0) 1 (0.3) 1 (0.7) 0 (0.0)

Muscle pain 9 (5.7) 40 (13.1) 16 (10.3) 7 (5.3) 20 (6.4) 22 (14.4) 7 (4.6)Severe 1 (0.6) 0 (0.0) 1 (0.6) 0 (0.0) 1 (0.3) 1 (0.7) 0 (0.0)

Oral temperature 1 (0.6) 2 (0.7) 2 (1.3) 0 (0.0) 0 (0.0) 0 (0.0) 0 (0.0)Severe 0 (0.0) 0 (0.0) 0 (0.0) 0 (0.0) 0 (0.0) 0 (0.0) 0 (0.0)

Gastrointestinal Systemic AEsDiarrhea 12 (7.6) 13 (4.3) 9 (5.8) 5 (3.8) 15 (4.8) 6 (3.9) 15 (9.9)

Severe 1 (0.6) 4 (1.3) 1 (0.6) 1 (0.8) 0 (0.0) 0 (0.0) 1 (0.7)Nausea 5 (3.2) 13 (4.3) 5 (3.2) 0 (0.0) 11 (3.5) 7 (4.6) 3 (2.0)

Severe 0 (0.0) 1 (0.3) 2 (1.3) 0 (0.0) 0 (0.0) 1 (0.7) 0 (0.0)Vomiting 0 (0.0) 5 (1.6) 1 (0.6) 0 (0.0) 1 (0.3) 1 (0.7) 1 (0.7)

Severe 0 (0.0) 1 (0.3) 1 (0.6) 0 (0.0) 0 (0.0) 1 (0.7) 0 (0.0)Respiratory/Facial AEs

Chest tightness 1 (0.6) 3 (1.0) 4 (2.6) 5 (3.8) 3 (1.0) 1 (0.7) 2 (1.3)Severe 0 (0.0) 0 (0.0) 0 (0.0) 0 (0.0) 1 (0.3) 0 (0.0) 0 (0.0)

Cough 11 (7.0) 12 (3.9) 12 (7.7) 5 (3.8) 10 (3.2) 3 (2.0) 8 (5.3)Severe 0 (0.0) 1 (0.3) 0 (0.0) 0 (0.0) 1 (0.3) 0 (0.0) 1 (0.7)

Difficulty breathing

0 (0.0) 3 (1.0) 3 (1.9) 1 (0.8) 4 (1.3) 3 (2.0) 1 (0.7)

Severe 0 (0.0) 0 (0.0) 0 (0.0) 0 (0.0) 1 (0.3) 0 (0.0) 1 (0.7)Difficulty swallowing

2 (1.3) 1 (0.3) 4 (2.6) 0 (0.0) 5 (1.6) 1 (0.7) 2 (1.3)

33

Severe 0 (0.0) 0 (0.0) 0 (0.0) 0 (0.0) 1 (0.3) 0 (0.0) 1 (0.7)Eye redness 3 (1.9) 1 (0.3) 1 (0.6) 1 (0.8) 6 (1.9) 1 (0.7) 1 (0.7)

Severe 0 (0.0) 0 (0.0) 1 (0.6) 0 (0.0) 0 (0.0) 0 (0.0) 0 (0.0)Eyelid swelling 2 (1.3) 2 (0.7) 0 (0.0) 0 (0.0) 2 (0.6) 0 (0.0) 0 (0.0)

Severe 0 (0.0) 0 (0.0) 0 (0.0) 0 (0.0) 0 (0.0) 0 (0.0) 0 (0.0)Facial swelling 0 (0.0) 1 (0.3) 0 (0.0) 0 (0.0) 1 (0.3) 0 (0.0) 0 (0.0)

Severe 0 (0.0) 0 (0.0) 0 (0.0) 0 (0.0) 0 (0.0) 0 (0.0) 0 (0.0)Hoarseness 5 (3.2) 10 (3.3) 4 (2.6) 1 (0.8) 8 (2.6) 3 (2.0) 1 (0.7)

Severe 0 (0.0) 1 (0.3) 0 (0.0) 0 (0.0) 1 (0.3) 0 (0.0) 0 (0.0)Sore throat 4 (2.5) 11 (3.6) 12 (7.7) 2 (1.5) 11 (3.5) 6 (3.9) 5 (3.3)

Severe 0 (0.0) 0 (0.0) 2 (1.3) 0 (0.0) 1 (0.3) 0 (0.0) 1 (0.7)Wheezing 1 (0.6) 3 (1.0) 3 (1.9) 0 (0.0) 3 (1.0) 3 (2.0) 3 (2.0)

Severe 0 (0.0) 0 (0.0) 0 (0.0) 0 (0.0) 1 (0.3) 0 (0.0) 0 (0.0)Abbreviations: AE, adverse event; HA, hemagglutinin; IIV3-HD, trivalent high-dose inactivated influenza vaccine; N, number

of subjects who received test article at Day 0; n, number of subjects in each specified category of adverse events; % =

(n/N)*100; qNIV, quadrivalent recombinant nanoparticle influenza vaccine; RIV4, quadrivalent recombinant influenza

vaccine.

Note: Percentages are based on the number of subjects in each treatment group who received test article on Day 0.

Treatment group in safety population is based on the actual dose(s) received. Subjects with multiple occurrences of the

same event were counted once for that event.

Note: Includes solicited adverse events reported by subjects (via diary or spontaneously) with a recorded start date within

the 7-day post-vaccination window (ie, from Study Day 0 to Study Day 6).

Note: For fever: mild = oral temperature 38.0-38.4°C, moderate = oral temperature 38.5-38.9 °C, severe = oral

temperature >38.9°C.

34

Table S7. Summary of Severe Adverse Events (SAEs) From Day 0 Post-vaccination to Day 182, Inclusive, by System Organ Class and Preferred Term – Safety Population

MedDRA System Organ Class and Preferred Term

Pooled Adjuvanted qNIV Groups

(Group A,B,C,D)

N=750n (%)

qNIV Unadjuvanted(Group E)

N=311n (%)

IIV3-HD(Group F)

N=153n (%)

RIV4(Group G)

N=151n (%)

Subjects with at least one SAE

44 (5.9) 6 (1.9) 6 (3.9) 3 (2.0)

Infections and infestations

12 (1.6) 1 (0.3) 2 (1.3) 1 (0.7)

Pneumonia 3 (0.4) 0 (0.0) 1 (0.7) 1 (0.7)

Diverticulitis 3 (0.4) 0 (0.0) 0 (0.0) 0 (0.0)

Sepsis 1 (0.1) 1 (0.3) 0 (0.0) 1 (0.7)

Cellulitis 1 (0.1) 1 (0.3) 0 (0.0) 0 (0.0)

Appendicitis 1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

Bronchitis 0 (0.0) 0 (0.0) 1 (0.7) 0 (0.0)

Bronchitis viral 1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

Clostridium difficile infection

1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

Influenza 1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

Staphylococcal infection

1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

Streptococcal sepsis

0 (0.0) 0 (0.0) 1 (0.7) 0 (0.0)

Urinary tract infection

1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

Nervous system disorders

8 (1.1) 0 (0.0) 0 (0.0) 0 (0.0)

Cerebrovascular accident

2 (0.3) 0 (0.0) 0 (0.0) 0 (0.0)

Seizure 2 (0.3) 0 (0.0) 0 (0.0) 0 (0.0)

Cerebral arteriosclerosis

1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

Dementia Alzheimer's type

1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

Embolic stroke 1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

Intracranial aneurysm

1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

35

Ischemic cerebral infarction

1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

Syncope 1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

Cardiac disorders 5 (0.7) 1 (0.3) 1 (0.7) 0 (0.0)

Acute myocardial infarction

1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

Angina pectoris 0 (0.0) 0 (0.0) 1 (0.7) 0 (0.0)

Angina unstable 1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

Atrial fibrillation 1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

Cardiac failure congestive

1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

Coronary artery disease

0 (0.0) 1 (0.3) 0 (0.0) 0 (0.0)

Myocardial infarction

1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

Gastrointestinal disorders

6 (0.8) 1 (0.3) 0 (0.0) 0 (0.0)

Gastrointestinal hemorrhage

2 (0.3) 0 (0.0) 0 (0.0) 0 (0.0)

Abdominal pain upper

1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

Duodenitis 1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

Dysphagia 1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

Gastroesophageal reflux disease

1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

Gastroptosis 1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

Hiatus hernia 1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

Intestinal perforation

1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

Esophageal stenosis

1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

Pancreatitis 0 (0.0) 1 (0.3) 0 (0.0) 0 (0.0)

Small intestinal obstruction

1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

Injury, poisoning, and procedural complications

2 (0.3) 1 (0.3) 1 (0.7) 2 (1.3)

Cervical vertebral fracture

0 (0.0) 0 (0.0) 0 (0.0) 1 (0.7)

Clavicle fracture 0 (0.0) 0 (0.0) 0 (0.0) 1 (0.7)

Femur fracture 1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

36

Head injury 0 (0.0) 0 (0.0) 1 (0.7) 0 (0.0)

Ligament injury 0 (0.0) 0 (0.0) 0 (0.0) 1 (0.7)

Pneumothorax traumatic

1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

Road traffic accident

0 (0.0) 0 (0.0) 0 (0.0) 1 (0.7)

Scapula fracture 1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

Spinal column injury

1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

Spinal fracture 1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

Upper limb fracture

0 (0.0) 1 (0.3) 0 (0.0) 0 (0.0)

Metabolism and nutrition disorders

3 (0.4) 0 (0.0) 2 (1.3) 0 (0.0)

Dehydration 2 (0.3) 0 (0.0) 1 (0.7) 0 (0.0)

Diabetic ketoacidosis

1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

Failure to thrive 1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

Hypokalemia 1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

Type 2 diabetes mellitus

0 (0.0) 0 (0.0) 1 (0.7) 0 (0.0)

Neoplasms benign, malignant and unspecified (incl cysts and polyps)

4 (0.5) 1 (0.3) 0 (0.0) 0 (0.0)

Breast cancer 1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

Lung cancer metastatic

1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

Malignant melanoma

1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

Non-small cell lung cancer metastatic

1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

Squamous cell carcinoma of the tongue

0 (0.0) 1 (0.3) 0 (0.0) 0 (0.0)

Respiratory, thoracic, and mediastinal disorders

4 (0.5) 0 (0.0) 1 (0.7) 0 (0.0)

Chronic obstructive pulmonary disease

1 (0.1) 0 (0.0) 1 (0.7) 0 (0.0)

37

Respiratory failure 2 (0.3) 0 (0.0) 0 (0.0) 0 (0.0)

Acute respiratory failure

1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

Pulmonary embolism

1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

General disorders and administration site conditions

3 (0.4) 0 (0.0) 0 (0.0) 0 (0.0)

Chest discomfort 1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

Chest pain 1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

Death 1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

Musculoskeletal and connective tissue disorders

2 (0.3) 1 (0.3) 0 (0.0) 0 (0.0)

Osteoarthritis 1 (0.1) 1 (0.3) 0 (0.0) 0 (0.0)

Spinal pain 1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

Blood and lymphatic system disorders

1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

Anemia 1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

Hepatobiliary disorders

1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

Cholelithiasis 1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

Hepatic cyst 1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

Renal and urinary disorders

1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

Calculus urinary 1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

Reproductive system and breast disorders

1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

Benign prostatic hyperplasia

1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

Skin and subcutaneous tissue disorders

1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

Vascular disorders 1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

Angioedema 1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

Aortic aneurysm 1 (0.1) 0 (0.0) 0 (0.0) 0 (0.0)

38

rupture

Abbreviations: AE, adverse event; N, number of subjects who receive test article at Day 0; IIV3-HD, trivalent high-dose

inactivated influenza vaccine; n = number of subjects in each specified category of MedDRA System Organ Class Preferred Term;

% = (n/N)*100; SAE, serious adverse event; qNIV, quadrivalent recombinant nanoparticle influenza vaccine; RIV4, quadrivalent

recombinant influenza vaccine.

Note: Treatment group in safety population is based on the actual dose(s) received. A subject who experienced multiple events

within a system organ class (SOC) was counted once. A subject who experienced multiple events within a preferred term was

counted once for that preferred term.

39

3 REFERENCES

1. Smith G, Liu Y, Flyer D, et al. Novel hemagglutinin nanoparticle influenza vaccine with Matrix-M™ adjuvant induces hemagglutination inhibition, neutralizing, and protective responses in ferrets against homologous and drifted A(H3N2) subtypes. Vaccine 2017; 35(40): 5366-72.

2. Lövgren Bengtsson K, Morein B, Osterhaus AD. ISCOM technology-based Matrix M™ adjuvant: success in future vaccines relies on formulation. Expert Rev Vaccines 2011; 10(4): 401-3.

3. Fluzone® High-Dose (Trivalent) Influenza Vaccine [package insert]. Swiftwater, PA: Sanofi Pasteur Inc.; 2019. Available at: https://www.fda.gov/media/119870/download . Accessed May 7, 2020.

4. Flublok® Quadrivalent Influenza Vaccine [package insert]. Meriden, CT: Protein Sciences Corporation; 2019.

5. World Health Organization. Recommended composition of influenza virus vaccines for use in the 2018-2019 northern hemisphere influenza season. February 22, 2018. Available at: https://www.who.int/influenza/vaccines/virus/recommendations/2018_19_north/en/. Accessed May 5, 2020.

6. Zost SJ, Parkhouse K, Gumina ME, et al. Contemporary H3N2 influenza viruses have a glycosylation site that alters binding of antibodies elicited by egg-adapted vaccine strains. Proc Natl Acad SciU S A 2017; 114(47): 12578-83.

7. Shinde V, Fries L, Wu Y, et al. Improved titers against influenza drift variants with a nanoparticle vaccine. N Engl J Med 2018; 378(24): 2346-8.

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https://www.who.int/influenza/vaccines/virus/recommendations/2018_19_north/en/

https://www.fda.gov/media/119870/download

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