Vaccine Information: EVUSHELD (Page 5 of 8)

12.4 Microbiology

Antiviral Activity

In a neutralization assay on Vero E6 cells, tixagevimab, cilgavimab, and their combination neutralized SARS-CoV-2 (USA-WA1/2020 isolate) with EC₅₀ values of 60.7 pM (9 ng/mL), 211.5 pM (32 ng/mL), and 65.9 pM (10 ng/mL), respectively.

Tixagevimab, cilgavimab, and their combination showed reduced or no antibody-dependent cell-mediated cytotoxicity (ADCC), antibody-dependent cellular phagocytosis (ADCP), or antibody-dependent natural killer cell activation (ADNKA) in cell culture studies. Tixagevimab, cilgavimab, and their combination did not mediate antibody-dependent complement deposition (ADCD) activity with guinea pig complement proteins.

Antibody Dependent Enhancement (ADE) of Infection

The potential of tixagevimab and cilgavimab to mediate antibody-dependent viral entry was assessed in FcγRII-expressing Raji cells co-incubated with recombinant virus-like particles (VLPs) pseudotyped with SARS-CoV-2 spike protein, with antibody concentrations at a range of 6.6 nM (1 µg/mL) to 824 pM (125 ng/mL). Tixagevimab, cilgavimab, and their combination did not mediate entry of VLPs into these cells under the tested conditions.

The potential for ADE was also evaluated in a non-human primate model of SARS-CoV-2 using EVUSHELD. Intravascular administration prior to virus inoculation resulted in a dose-dependent improvement in all measured outcomes (total viral RNA in the lungs or nasal mucosae, infectious virus levels in the lungs based on TCID₅₀ measurements, or lung injury and pathology based on histology measurements). No evidence of enhancement of viral replication or disease was observed at any dose evaluated, including sub-neutralizing doses down to 0.04 mg/kg.

Antiviral Resistance

There is a potential risk of treatment failure due to the development of viral variants that are resistant to tixagevimab and cilgavimab. Prescribing healthcare providers should consider the prevalence of SARS-CoV-2 variants in their area, where data are available, when considering prophylactic treatment options.

Escape variants were identified following serial passage in cell culture of SARS-CoV-2 or replication competent recombinant vesicular stomatitis virus (VSV) expressing SARS-CoV-2 spike protein in the presence of tixagevimab or cilgavimab individually or in combination. Tixagevimab selected a variant expressing F486S in the spike protein with a >800-fold reduction in susceptibility to tixagevimab. Cilgavimab selected variants that expressed spike protein amino acid substitutions R346G, R346I, K444E, K444N, K444Q, K444R, K444T or N450D were each associated with a >200-fold reduction in susceptibility to cilgavimab. No escape variants to the tixagevimab and cilgavimab combination were selected.

In neutralization assays using recombinant VLPs pseudotyped with SARS-CoV-2 spike and harboring individual spike amino acid substitutions identified in circulating SARS-CoV-2, variants with reduced susceptibility to cilgavimab alone included those with R346I (>200-fold), K444E (>200-fold), K444Q (>200-fold), K444R (>200-fold), V445A (21- to 51-fold), G446V (4.2-fold), N450K (9.1-fold), or L452R (5.8-fold) substitutions. Variants with reduced susceptibility to tixagevimab alone included those with Q414R (4.6-fold), L455F (2.5- to 4.7-fold), G476S (3.3-fold), E484D (7.1-fold), E484K (6.2- to 12-fold), E484Q (3.0-fold), F486S (>600-fold), F486V (121- to 149-fold), Q493K (2.4- to 3.2-fold), Q493R (7.9-fold), E990A (6.1-fold), or T1009I (8.2-fold) substitutions. Variants harboring an E484K (2.4- to 5.4-fold), Q493R (3.4-fold), E990A (5.7-fold), or T1009I (4.5-fold) substitution exhibited low level reduced susceptibility to tixagevimab and cilgavimab in combination.

VLPs pseudotyped with the SARS-CoV-2 spike of variant strains with reduced susceptibility to cilgavimab included those with R346K+E484K+N501Y (Mu, 21-fold), and those with reduced susceptibility to tixagevimab included those harboring E484K (Alpha, 18.5-fold; Beta, 3.5- to 15-fold; Zeta, 7.3-fold). Similar results were observed, where data were available, in neutralization assays using authentic SARS-CoV-2 variant strains.

VLPs pseudotyped with the SARS-CoV-2 spike of Omicron BA.1 or BA.1.1 (BA.1+R346K) showed reduced susceptibility to tixagevimab (>600- to >1,000-fold or 460-fold, respectively) and to cilgavimab (>700- to >1,000-fold or >500-fold, respectively). VLPs pseudotyped with the SARS-CoV-2 spike of Omicron BA.2 or BA.2.12.1 showed reduced susceptibility to tixagevimab (>1,000-fold or >500-fold, respectively) but not to cilgavimab (1.9-fold or 2-fold, respectively). VLPs pseudotyped with the SARS-CoV-2 spike of Omicron BA.2.75 or BA.2.75.2 showed reduced susceptibility to tixagevimab (7- to 53-fold or >3,333- to >10,000-fold, respectively) and to cilgavimab (6- to 40-fold or >769- to >5,000-fold, respectively). VLPs pseudotyped with the SARS-CoV-2 spike of Omicron BA.3 showed reduced susceptibility to tixagevimab (>5,000-fold) but not to cilgavimab (4-fold). VLPs pseudotyped with the SARS-CoV-2 spike of Omicron BA.4/BA.5 showed reduced susceptibility to tixagevimab (>10,000-fold) and cilgavimab (7.5- to 9-fold). VLPs pseudotyped with the SARS-CoV-2 spike of Omicron BA.4.6 showed reduced susceptibility to tixagevimab (>1,000-fold) and to cilgavimab (>1,000-fold). VLPs pseudotyped with the SARS-CoV-2 spike of Omicron BF.7 or BJ.1 showed reduced susceptibility to tixagevimab (>3,333- to >10,000-fold or 85- to 172-fold, respectively) and to cilgavimab (>769- to >5,000-fold or >769- to >5,000-fold, respectively). VLPs pseudotyped with the SARS-CoV-2 spike of Omicron BQ.1 or BQ.1.1 showed reduced susceptibility to tixagevimab (>1,250- to >10,000-fold) and to cilgavimab (>667- to >5,000-fold). VLPs pseudotyped with the SARS-CoV-2 spike of Omicron BQ.1 or BQ.1.1 showed reduced susceptibility to tixagevimab (>1,250- to >10,000-fold) and to cilgavimab (>667- to >5,000-fold). VLPs pseudotyped with the SARS-CoV-2 spike of Omicron BA.5.2.6 or BF.11 showed reduced susceptibility to tixagevimab (>333-fold) and to cilgavimab (>77-fold). VLPs pseudotyped with the SARS-CoV-2 spike of Omicron BN.1 or XBB showed reduced susceptibility to tixagevimab (24- to 44-fold or >2,600- to >10,000-fold, respectively) and to cilgavimab (>3,700- to >5,000-fold or >565- to >5,000-fold, respectively). The effects of the individual substitutions in Omicron spike glycoproteins on neutralization susceptibility are being investigated.

The neutralizing activity of tixagevimab and cilgavimab in combination was tested against pseudotyped VLPs and/or authentic SARS-CoV-2 variant strains harboring all spike substitutions identified in Alpha (B.1.1.7, 0.5- to 5.2-fold), Beta (B.1.351, 1.0- to 3.8-fold), Gamma (P.1, 0.4- to 2.0-fold), Delta (B.1.617.2, 0.6- to 1.2-fold), and Delta [+K417N] (AY.1/ AY.2, 1.0-fold) variants of concern, and Eta (B.1.525, 3.1-fold), Iota (B.1.526, 0.3- to 3.4-fold), Kappa (B.1.617.1, 0.5- to 3.4-fold) Lambda (C.37, 0.7-fold), and Mu (B.1.621, 7.5-fold) variants of interest. Tixagevimab and cilgavimab in combination was also tested against Epsilon (B.1.427 / B.1.429, 0.8- to 3.5-fold), R.1 (3.5-fold), B.1.1.519 (1.4-fold), C.36.3 (2.3-fold), B.1.214.2 (0.8-fold), and B.1.619.1 (3.3-fold) variant alerts for further monitoring and B.1.616 (0.5 fold), A.23.1 (0.4-fold), A.27 (0.8-fold), and AV.1 (5.9-fold) variants de-escalated from further monitoring (Table 6).

Preliminary data for the neutralizing activities of tixagevimab and cilgavimab in combination against circulating Omicron subvariants are available. VLPs pseudotyped with the SARS-CoV-2 spike of Omicron BA.1 or BA.1.1 (BA.1+R346K) showed reduced neutralizing activity (132- to 183-fold or 424-fold, respectively), Omicron BA.2 showed no change in neutralizing activity (3.2-fold). VLPs pseudotyped with the spike of Omicron BA.2.12.1, BA.2.75, BA.2.75.2, BA.3, BA.4/BA.5, or BA.4.6 showed 5-fold, 2.4- to 15-fold, >5,000- to >10,000-fold, 16-fold, 33- to 65-fold, or >1,000-fold reductions in neutralizing activity, respectively. VLPs pseudotyped with the spike of Omicron BF.7, BJ.1, BQ.1 or BQ.1.1 showed >5,000- to >10,000-fold, 228- to 424-fold, >2,000- to >10,000-fold or >2,000- to >10,000-fold reductions in neutralizing activity, respectively. VLPs pseudotyped with the spike of Omicron BA.5.2.6, BF.11, BN.1 or XBB showed >500-fold, >500-fold, 68-fold or >1,400- to >10,000-fold reductions in neutralizing activity, respectively. Authentic Omicron BA.1, BA.1.1, BA.2, or BA.5 viruses showed 12- to 30-fold, 176-fold, 5.4-fold, or 2.8- to 16-fold reductions in susceptibility, respectively.

Data collection is ongoing to better understand how the reductions in activity seen in pseudotyped VLP assays or authentic SARS-CoV-2 assays may correlate with clinical outcomes. Emerging Omicron subvariants that are resistant to neutralization by cilgavimab harbor the spike substitution R346T or K444T, while those resistant to neutralization by tixagevimab harbor the spike substitution F486S or F486V. EVUSHELD is unlikely to neutralize SARS-CoV-2 Omicron subvariants harboring R346T or K444T in combination with F486S or F486V.

Table 6 EVUSHELD Pseudotyped Virus-Like Particles and Authentic SARS-CoV-2 Neutralization Data for SARS-CoV-2 Variants

Lineage with Spike Protein Substitution	Country First Identified	WHO Nomenclature	Key Substitutions Tested	Fold Reduction in Susceptibility * (Pseudotyped VLPs † )	Fold Reduction in Susceptibility * (Authentic virus ‡ )
* Range of reduced potency across multiple variants of each lineage using research-grade pseudotyped VLP neutralization assays; mean fold change in half maximal effective concentration (EC50 ) of mAb required for a 50% reduction in infection compared to wild type reference strain † Pseudotyped virus-like particles expressing the entire SARS-CoV-2 spike variant protein and individual characteristic spike substitutions except L452Q were tested including Alpha (+L455F, E484K, F490S, Q493R, and/or S494P), and Delta (+K417N) harboring additional indicated RBD substitutions that are no longer detected or detected at extremely low levels within these lineages ‡ Authentic SARS-CoV-2 expressing the entire variant spike protein were tested including Alpha (+E484K or S494P) harboring additional indicated RBD substitutions that are no longer detected or detected at extremely low levels within these lineages § No change: <5-fold reduction in susceptibility ¶ EC50 value = 1.13 – 1.83 nM (171 — 277 ng/mL) # Tixagevimab and cilgavimab together are unlikely to be active against this variant.
B.1.1.7	UK	Alpha	N501Y	0.5- to 5.2‑fold	No Change §
B.1.351	South Africa	Beta	K417N+E484K+N501Y	No Change §	No Change §
P.1	Brazil	Gamma	K417T+E484K+N501Y	No Change §	No Change §
B.1.617.2	India	Delta	L452R+T478K	No Change §	No Change §
AY.1/ AY.2	India	Delta [+K417N]	K417N+L452R+T478K	No Change §	No Change §
BA.1	Botswana	Omicron (BA.1)	G339D+S371L+S373P+ S375F+K417N+N440K+ G446S+S477N+T478K+ E484A+Q493R+G496S+ Q489R+N501Y+Y505H	132- to 183-fold ¶	12- to 30-fold
BA.1.1	Multiple country origin	Omicron (BA.1.1) [+R346K]	BA.1+R346K	424-fold	176-fold
BA.2	Multiple country origin	Omicron (BA.2)	G339D+S371F+S373P+ S375F+T376A+D405N+ R408S+K417N+N440K+ S477N+T478K+E484A+ Q493R+Q498R+N501Y+ Y505H	No Change §	5.4-fold
BA.2.12.1	United States	Omicron (BA.2.12.1)	BA.2+L452Q	5-fold	ND
BA.2.7.5	India	Omicron (BA.2.75)	G339H+S371F+S373P+ S375F+T376A+D405N+ R408S+K417N+N440K+ G446S+N460K+S477N+ T478K+E484A+Q498R+ N501Y+ Y505H	2.4- to 15-fold	ND
BA.2.75.2	India	Omicron (BA.2.75.2)	BA.2.75+R346T+F486S	>5000-fold #	ND
BA.3	Mulitple country origin	Omicron (BA.3)	G339D+S371F+ S373P+S375F+D405N+ K417N+N440K+G446S+ S477N+T478K+E484A+ Q493R+Q498R+N501Y+ Y505H	16-fold	ND
BA.4	Mulitple country origin	Omicron (BA.4)	G339D+S371F+S373P+ S375F+T376A+D405N+ R408S+K417N+N440K+ L452R+S477N+T478K+ E484A+F486V+Q498R+ N501Y+Y505H	33- to 65-fold	ND
BA.4.6	United States	Omicron (BA.4.6)	BA.4+R346T	>1000-fold #	ND
BA.5	Mulitple country origin	Omicron (BA.5)	+G339D+S371F+S373P+ S375F+T376A+D405N+ R408S+K417N+N440K+ L452R+S477N+T478K+ E484A+F486V+Q498R+ N501Y+Y505H	33- to 65-fold	2.8- to 16-fold
BA.5.2.6	Mulitple country origin	Omicron (BA.5.2.6)	G339D+R346T+S371F+ S373P+S375F+T376A+ D405N+R408S+K417N+ N440K+L452R+S477N+ T478K+E484A+F486V+ Q498R+N501Y+Y505H	>500-fold	ND
BF.7	United States/Belgium	Omicron (BF.7)	BA.4+R346T	>5000-fold #	ND
BF.11	Mulitple country origin	Omicron (BF.11)	G339D+R346T+S371F+ S373P+ S375F+T376A+ D405N+R408S+K417N+ N440K+L452R+S477N+ T478K+E484A+F486V+ Q498R+N501Y+Y505H	>500-fold	ND
BJ.1	Mulitple country origin	Omicron (BJ.1)	G339H+R346T+L368I+ S371F+S373+S375F+ T376A+D405N+R408S+ K417N+N440K+V445P+ G446S+S477N+T478K+ V483A+E484A+F490V+ Q493R+Q498R+N501Y+ Y505H	228- to 424-fold	ND
BN.1	Mulitple country origin	Omicron (BN.1)	G339D+R346T+K356T+ S371F+S373P+S375F+ D405N+ R408S+ K417N+N440K+G446S+ N460K+S477N+T478K+ E484A+F490S+ Q493R+Q498R+Y505H	68-fold	ND
BQ.1	Nigeria	Omicron (BQ.1)	BA.5+K444T+N460K	>2000-fold #	ND
BQ.1.1	Multiple country origin	Omicron (BQ.1.1)	BA.5+R346T+K444T+ N460K	>2000-fold #	ND
XBB	Multiple country origin	Omicron (XBB)	G339H+R346T+L368I+ S371F+S373P+S375F+ T376A+D405N+R408S+ K417N+N440K+V445P+ G446S+N460K+S477N+ T478K+ E484A+F486S+ F490S+Q498R+N501Y+ Y505H	>1400-fold #	ND
B.1.525	Multiple country origin	Eta	E484K	No Change §	ND
B.1.526	United States	Iota	E484K	No Change §	No Change §
B.1.617.1	India	Kappa	L452R+E484Q	No Change §	No Change §
C.37	Peru	Lambda	L452Q+F490S	No Change §	ND
B.1.621	Colombia	Mu	R346K+E484K +N501Y	7.5-fold	ND
B.1.427 / B.1.429	United States	Epsilon	L452R	No Change §	No Change §
R.1	Multiple country origin	–	E484K	No Change §	ND
B.1.1.519	Multiple country origin	–	T478K	No Change §	ND
C.36.3	Multiple country origin	–	R346S:L452R	No Change §	ND
B.1.214.2	Multiple country origin	–	Q414K:N450K	No Change §	ND
B.1.619.1	Multiple country origin	–	N440K:E484K	No Change §	ND
P.2	Brazil	Zeta	E484K	No Change §	ND
B.1.616	France	–	V483A	No Change §	ND
A.23.1	UK	–	V367F	No Change §	ND
A.27	Multiple country origin	–	L452R+N501Y	No Change §	ND
AV.1	Multiple country origin	–	N439K+E484K	5.9-fold	ND
ND, not determined; RBD, receptor binding domain

It is not known how pseudotyped VLPs or authentic SARS-CoV-2 neutralization susceptibility data correlate with clinical outcome.

In PROVENT, illness visit sequencing data were available for 21 of 33 subjects with SARS-CoV-2 infection (6 who received tixagevimab and cilgavimab and 15 placebo). Fourteen subjects were infected with variants of concern or variants of interest, including 8 subjects with Alpha (B.1.1.7) (8 who received placebo), 1 subject with Beta (B.1.351) (1 who received tixagevimab and cilgavimab), 3 subjects with Delta (B.1.617.2) (3 who received placebo), and 2 subjects with Epsilon (B.1.429) (2 who received tixagevimab and cilgavimab). Seven additional subjects were infected with B.1.375 (1 who received tixagevimab and cilgavimab) or the A_1 set of lineages containing a constellation of spike protein substitutions including D614G and P681H or Q677P (3 who received tixagevimab and cilgavimab and 3 placebo). Additional spike protein RBD substitutions detected at low frequency (between 3% and 24%) included V503F in the tixagevimab and cilgavimab group.

In STORM CHASER, illness visit sequencing data was available for 19 of 19 subjects with SARS-CoV-2 infections (12 of 12 who received tixagevimab and cilgavimab and 7 of 7 placebo). At an allele fraction ≥25%, 12 of 19 subjects were infected with variants of concern or variants of interest, including 9 subjects with Alpha (B.1.1.7) (5 who received tixagevimab and cilgavimab and 4 placebo) and 3 subjects with Epsilon (B.1.427 / B.1.429) (2 who received tixagevimab and cilgavimab and 1 placebo). Seven additional subjects were infected with B.1.1.519 (1 who received tixagevimab and cilgavimab) or the A_1 set of lineages containing a constellation of spike protein substitutions including D614G and D138H, Q675H, Q677H, or V1176F (4 who received tixagevimab and cilgavimab and 2 placebo). Additional spike protein RBD substitutions detected at an allele fraction ≥3% included S325P, Del342, C361W, Del428, F429V, and F515C in the tixagevimab and cilgavimab group.

Evaluation of neutralization susceptibility of variants identified through global surveillance and in subjects who received tixagevimab and cilgavimab is ongoing.

It is possible that variants resistant to tixagevimab and cilgavimab could have cross-resistance to other monoclonal antibodies targeting the RBD of SARS-CoV-2. The combination of tixagevimab and cilgavimab retained activity against pseudotyped VLPs harboring individual SARS-CoV-2 spike substitutions (K417E/N, D420N, K444Q, V445A, Y453F, L455F, N460K/S/T, E484D/K/Q, F486V, F490S, Q493K/R, and S494P) identified in neutralization escape variants of other monoclonal antibodies targeting the RBD of SARS-CoV-2 spike protein.