8 th Vaccine & ISV Congress October 2014 Philadelphia, USA

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1 8 th Vaccine & ISV Congress October 2014 Philadelphia, USA 2014 Guinea Ebola Virus Recombinant Glycoprotein (GP) Nanoparticle Vaccine was Highly Immunogenic and Cross- Neutralized 1976 Ebola Virus GP Pseudovirus Gale Smith 1, Ye Liu 1, David Flyer 1, Louis Fries 1, Karin Lovgren 2, Jay Hooper 3, and Gregory Glenn 1 1 Novavax Inc., Gaithersburg, MD 2 Novavax AB, Uppsala, Sweden 3 USAMRIID, Fort Detrick, MD

2 In memoriam: Tragically, five co-authors, who contributed greatly to public health and research efforts in Sierra Leone, contracted EVD and lost their battle with the disease before this manuscript could be published In Sierra Leone May 2014 at funeral of a Guinea EVD case, attendees were infected with two distinct Ebola cluster 1,2 viruses leading to cluster 3 sustained human-to-human transmission. Because many of the mutations alter protein sequences and other biologically meaningful targets, they should be monitored for impact on diagnostics, vaccines, and therapies critical to outbreak response. 2

3 Ebola GP Sequence Comparison Mayinga 1976 (Discovered) vs Guinea 2014 (Epidemic) 1976 GP 2014 GP: 20 AA mutations mab 13F6 Lee et al, Nature (7201):

4 Novavax EBOV [H.sapiens-wt/SLE/2014/ManoRiver G3798, cluster 3] Glycoprotein (GP) Nanoparticle Vaccine EBOV GP based on the Guinea, cluster 3 gene sequence; Gire, et al Science;12 September 2014 Full length, unmodified EBOV GP 1,2 [H.sapiens -wt/sle/2014/ ManoRiver-G3798; cluster 3] Clone genes into baculovirus Infect Sf9 insect cells with baculovirus Sf9 cells express protein antigens EBOV GP 4 4

5 5 Purified 2014 Guinea Ebola Recombinant GP

6 Capillary Electrophroesis (CE) Deglyosylated/Reduced Ebola GP 1,2 Nanoparticles PDA - 220nm Ebola GP 1613 Red Degly Corrected Area Percent PDA - 220nm Ebola GP 1613 Reduced Corrected Area Percent AU AU 54.4 p10kd marker GP PNGase F GP Minutes 6

7 Guinea EBOV Recombinant GP 36.7nm Particles

8 EBOLA GP particle size 36.7 nm Size Distribution by Intensity 15 Intensity (%) Size (d.nm) Record 1: Ebola 08Sep14 Z-Ave (d.nm) Mean DLS PdI Mean PdI

9 Manufacturing Process Ebola GP Nanoparticles GMP Manufacture GP nanoparticles 200L 1,000 L scale Non-ionic detergent extraction GP Chromatography: IEX and Affinity Forms Ebola pre-fusion, glycosylated GP multioligomeric 36.7nm micelles (protein nanoparticles) Purity 95% 2 8C storage in buffered saline, no preservatives Nanoparticles manufacturing process RSV F nanoparticle vaccine Phase 1/2 Adults, pregnant women, elderly, children Rabies G nanoparticle vaccine Phase 1/2 in India 9

10 Human Neutralizing mab KZ52 Binds to Pre-fusion Ebola GP Ebola KZ52 Human Monoclonal Antibody* Isolated from a human survivor of 1995 outbreak in Kikwit, Zaire Binds residues at the N terminus of GP1, and and at the N terminus of GP2 Requires an epitope seen only in the GP2 pre-fusion conformation Protection from lethal EBOV challenge in rodents; minimal protection of non-human primates Lee et al, Nature (7201):

11 Binding mab KZ52 to recombinant, purified Ebola GP [H.sapiens - wt/sle/2014/ ManoRiver-G3798; cluster 3] Ebola KZ52 SRP Analysis* *Hood.C.L. et al (2010) J. Virol 84, RU Novavax Ebola GP nanoparticles NVAX ZEBOV-GP 1,2 k a =3.5e 4 M -1 s -1, k d =1.3e -5 s -1, K D =0.37nM Ligand Antigen K a (1/ M Sec) K d (1/sec) K D (nm) KZ52 GPdTM 4.5E E nm* KZ52 EBOL GP 1,2 (NVAX) 3.5E E nm 11

12 EBOV mab Affinity Recombinant GP Nanoparticles mab Epitope Neutralization Protection in vivo KZ52 aa 42-43, 513, , Conformational + GP C6 aa Conformational + NHP 6D8 aa HNTPVYKLDISEATQVE 13F6 aa ATQVEQHHRRTDNDSTA Linear + NHP Linear + NHP 12

13 Anti-Ebola ELISA EC 50 of mab 13C6, 6D8 and KZ Ebola mab ELISA KZ52, 13C6, 6D F6 0 1e-5 1e mab conccentration (ug/ml) EC 50 ( g/ml) 4-P Fit: y = (A - D)/( 1 + (x/c)^b ) + D: A B C D R^2 c13c6 (Group01: Dilution vs Values) H13F6 (Group02: Dilution vs Values) e e C c6d8 (Group03: Dilution vs Values) KZ52 (Group04: Dilution vs Values) F6 * - Weighting: Fixed 6D KZ aa change in 13F6 epitope on Ebola GP nanoparticle results in loss of recognition 13

14 Immunogenicity of Recombinant 2014 EBOV GP Vaccine in Mice STUDY DESIGN: Group N EBOLA Vaccine GP Dose (µg) Adjuvant (µg) Immun days Blood Draw days , 14 0, 21, Matrix M (5) 0, 14 0, 21, Al PO 4 (50) 0, 14 0, 21, Saline - - 0, 21, 28 14

15 Recombinant EBOV GP Vaccine Anti-GP ELISA (EC90) Responses in Mice

16 Cross-Neutralization 2014 (Guinea) EBOV GP Vaccine of 1976 (Mayinga) PsVNA50 Pseudovirus PsVNA

17 Characterization Guinea 2014 Recombinant EBOV GP A baculovirus was constructed to express full-length, unmodified EBOV GP gene [H.sapiens-wt/SLE/2014/ManoRiver-G3798; cluster 3]. Purified EBOV GP trimers were chalice or cup-like pre-fusion structures that self-assembled into 36.7nm multi-oligomeric protein particles. Human monoclonal antibody KZ52 bound with high affinity to the purified recombinant EBOV GP, confirming that recombinant GP trimers were in a native pre-fusion conformation and KZ52 neutralizing epitope was intact. 17

18 Recombinant 2014 EBOV GP Nanoparticle Vaccine Cross-Neutralization 1976 VSV-GP Pseudovirus One week after a second immunization in mice of 5µg 2014 EBOV GP vaccine, reciprocal antibody titers against the homologous 2014 Guinea EBOV GP were high, (ELISA EC 90 ). Adsorption to aluminum phosphate to recombinant GP provided only a modest adjuvant effect. However, Matrix-M saponin adjuvant enhanced anti-gp responses fold, inducing Ebola GP antibody titers of Most significantly, 2014 EBOV GP plus Matrix-M induced serum crossneutralization to unexpectedly high antibody titers (PsVNA50) of against the older Ebola 1976 (Mayinga) GP-VSV pseudotype. The 2014 Guinea EBOV GP nanoparticle vaccine plus Matrix-M induced anti GP ELISA and neutralizing antibody levels well within ranges reported to protect against challenge with Ebola viruses in rodent and non-human animal models. 18

19 Conclusions This is the first report of a EBOV GP vaccine candidate produced against an epidemic 2014 West African EBOV GP sequence. Recombinant GP was correctly folded, efficiently produced using a scaleable technology, highly purified, and formed nanoparticles that were highly immunogenic with Matrix-M adjuvant inducing cross-neutralizing antibodies. The Novavax Guinea 2014 recombinant Ebola GP nanoparticle vaccine should not require frozen storage and can be manufactured quickly and at large scale to potentially produce millions of vaccine doses. Safety, immunogenicity, and challenge studies including in non-human primates of the 2014 Guinea recombinant EBOV GP nanoparticle vaccine were initiated. Human trials are scheduled with and without Matrix-M adjuvant to evaluate the potential for dose-sparing and broadening of epitope responses, as recently demonstrated with avian influenza recombinant H7N9 vaccine human trials (Glenn, et al NEJM Nov /NEJMc ; unpublished). 19