Bebtelovimab: Package Insert and Label Information (Page 3 of 5)

12.4 Microbiology

Antiviral Activity

The cell culture neutralization activity of bebtelovimab against SARS-CoV-2 was measured in a dose-response model quantifying plaque reduction using cultured Vero E6 cells. Bebtelovimab neutralized the USA/WA/1/2020 isolate of SARS-CoV-2 with an estimated EC50 value = 0.044 nM (6.4 ng/mL).

Bebtelovimab demonstrated antibody-dependent cell-mediated cytotoxicity on Jurkat reporter cells expressing FcγRIIIa following engagement with target cells expressing spike protein. Bebtelovimab did not elicit complement-dependent cytotoxicity activity in cell-based assays.

Antibody Dependent Enhancement (ADE) of Infection

The risk that bebtelovimab could mediate viral uptake and replication by immune cells was studied in THP-1 and Raji cell lines and primary human macrophages. In general, experiments with bebtelovimab did not demonstrate productive viral infection in immune cells exposed to SARS-CoV-2 at concentrations of mAb down to 60,000-fold below the approximate EC50 value for neutralization.

Antiviral Resistance

There is a potential risk of treatment failure due to the development of viral variants that are resistant to bebtelovimab.

Nonclinical selection studies using a directed evolution of a yeast displayed Spike RBD identified that substitutions at residues K444, V445, G446, and P499 interfered with bebtelovimab’s ability to block the Spike RBD:ACE-2 interaction. Pseudotyped virus-like particle (VLP) neutralization assays confirmed a 5-fold or greater reduction in susceptibility to bebtelovimab of viral variants with the following substitutions: K444E (>862), K444N (>1,901-fold), K444Q (208-fold), K444T (>1,814-fold), V445A (111-fold), V445F (369-fold), V445G (>730-fold), G446D (69-fold), G446R (7-fold), G446V (8-fold), P499H (>1,606-fold), P499R (>1,870-fold), and P499S (25-fold). In the context of Delta spike protein, G446V substitution had reduced susceptibility of 16.4-fold.

Pseudotyped VLP assessment using the full-length spike genes from different variant lineages indicate that bebtelovimab retains activity (<5-fold reduction) against the Alpha (B.1.1.7, UK origin), Beta (B.1.351, South Africa origin), Gamma (P.1, Brazil origin), Delta (B.1.617.2, India origin), Delta [+K417N] (AY.1/AY.2, India origin), Epsilon (B.1.427/B.1.429, California origin), Iota (B.1.526, New York origin), Kappa (B.1.617.1, India origin), Lambda (C.37, Peru origin), Omicron (B.1.1.529/BA.1, South Africa origin), Omicron [+R346K] (BA.1.1), Omicron BA.2, Omicron BA.2 [+L452Q] (BA.2.12.1), Omicron BA.2 [+D339H, G446S, N460K, R493Q (reversion)] (BA.2.75), and Omicron BA.4/BA.5 variant lineages (Table 2). The Mu (B.1.621, Colombia origin) variant showed a reduction in susceptibility to bebtelovimab of 5.3-fold.

Table 2: Bebtelovimab Pseudotyped Virus-Like Particle Neutralization Data for SARS-CoV-2 Spike Protein Variants

a Key substitutions occurring in the receptor binding domain of spike protein are listed. Pseudotyped VLP contained the full-length spike protein reflective of the consensus sequence for each of the variant lineages.

b No change: <5-fold reduction in susceptibility.

c Isolates of the B.1.526 lineage harbor several spike protein amino acid substitutions, and not all isolates contain the E484K substitution (as of February 2021).

Lineage with Spike Protein Substitution Country First Identified WHO Nomenclature Key Substitutions Tested a Fold Reduction in Susceptibility
B.1.1.7 UK Alpha N501Y No changeb
B.1.351 South Africa Beta K417N + E484K + N501Y No changeb
P.1 Brazil Gamma K417T + E484K + N501Y No changeb
B.1.617.2/AY.3 India Delta L452R + T478K No changeb
AY.1/AY.2(B.1.617.2 sublineages) India Delta [+K417N] L452R + T478K + K417N No changeb
B.1.427/B.1.429 USA (California) Epsilon L452R No changeb
B.1.526c USA (New York) Iota E484K No changeb
B.1.617.1 India Kappa L452R + E484Q No changeb
C.37 Peru Lambda L452Q + F490S No changeb
B.1.621 Colombia Mu R346K + E484K + N501Y 5.3
B.1.1.529/BA.1 South Africa Omicron [BA.1] G339D + S371L + S373P + S375F + K417N + N440K + G446S + S477N + T478K + E484A + Q493R + G496S + Q498R + N501Y + Y505H No changeb
BA.1.1 South Africa Omicron [+R346K] BA.1 + R346K No changeb
BA.2 South Africa Omicron [BA.2] G339D + S371F + S373P + S375F + T376A + D405N + R408S + K417N + N440K + S477N + T478K + E484A + Q493R + Q498R + N501Y + Y505H No changeb
BA.2.12.1 USA Omicron [BA.2+L452Q] BA.2 + L452Q No changeb
BA.2.75 India Omicron [BA.2+D339H, G446S, N460K, R493Q (reversion)] BA.2 + D339H, G446S, N460K, R493Q (reversion) No changeb
BA.4/BA.5 South Africa Omicron [BA.4/BA.5] G339D + S371F + S373P + S375F + T376A + D405N + R408S + K417N + N440K + L452R + S477N + T478K + E484A + F486V + Q498R + N501Y + Y505H No changeb

In authentic SARS-CoV-2 assays, bebtelovimab retained activity (<5-fold reduction) against variant virus isolates from the Alpha (B.1.1.7), Beta (B.1.351), Gamma (P.1), Delta (B.1.617.2/AY.3), Omicron (B.1.1.529/BA.1), Omicron [+R346K] (BA.1.1), Omicron BA.2, Omicron BA.2 [+L452Q] (BA.2.12.1), and Omicron BA.4 lineages, as well as SARS-CoV-2 (USA/WA/1/2020 isolate) engineered to express the L452R substitution present in the Epsilon (B.1.427/B.1.429) lineage or the E484K substitution present in the Iota (B.1.526) lineage (Table 3).

Table 3: Authentica SARS-CoV-2 Neutralization Data for Bebtelovimab

a The B.1.1.7, B.1.351, B.1.617.2, B.1.1.529/BA.1, and BA.2 variants were assessed using cell culture-expanded virus isolates and tested using a plaque reduction assay; the B.1.351, P.1, B.1.617.2, B.1.1.529/BA.1, BA.1.1, BA.2, BA.2.12.1, and BA.4 variants were assessed using cell culture-expanded isolates and tested using a microneutralization assay with a CPE-based endpoint titer to determine the IC>99 ; the B.1.526/E484K and B.1.427/B.1.429/L452R substitutions were assessed using recombinant SARS-CoV-2 (USA/WA/1/2020 isolate with E484K or L452R) and tested using a plaque reduction assay.

b Key substitutions occurring in receptor binding domain of spike protein which are associated with each lineage.

c No change: <5-fold reduction in susceptibility when compared to ancestral control isolate using the same methodology.

d These viral variants have been tested with two different neutralization methodologies, both yielding <5-fold reductions in susceptibility.

e Isolates of the B.1.526 lineage harbor several spike protein amino acid substitutions, and not all isolates contain the E484K substitution (as of February 2021).

Lineage with Spike Protein Substitution Country First Identified WHO Nomenclature Key Substitutions Tested b Fold Reduction in Susceptibility
B.1.1.7 UK Alpha N501Y No changec
B.1.351 South Africa Beta K417N, E484K, N501Y No changec,d
P.1 Brazil Gamma K417T, E484K, N501Y No changec
B.1.617.2/AY.3 India Delta L452R, T478K No changec,d
B.1.427/B.1.429 USA (California) Epsilon L452R No changec
B.1.526e USA (New York) Iota E484K No changec
B.1.1.529/BA.1 South Africa Omicron G339D + S371L + S373P + S375F + K417N + N440K + G446S + S477N + T478K + E484A + Q493R + G496S + Q498R + N501Y + Y505H No changec,d
BA.1.1 South Africa Omicron [+R346K] BA.1 + R346K No changec
BA.2 South Africa Omicron [BA.2] G339D + S371F + S373P + S375F + T376A + D405N + R408S + K417N + N440K + S477N + T478K + E484A + Q493R + Q498R + N501Y + Y505H No changec, d
BA.2.12.1 USA Omicron [BA.2+L452Q] BA.2 + L452Q No changec
BA.4 South Africa Omicron [BA.4] G339D + S371F + S373P + S375F + T376A + D405N + R408S + K417N + N440K + L452R + S477N + T478K + E484A + F486V + Q498R + N501Y + Y505H No changec

Genotypic analysis and phenotypic testing are ongoing to monitor for potential bebtelovimab-resistance-associated spike variations in clinical trials. Baseline sequencing data are available for 611 of the subjects in the BLAZE-4 (Arms 9-14) Study. Of these, 552 (90.3%) were infected with a variant of interest or concern, as designated by the WHO. No subject was infected with virus of the Omicron lineage or sub-lineages. The majority of subjects in the trial were infected with Delta (49.9%) and Alpha (28.6%). These were distributed across the treatment groups with Delta and Alpha infection rates of 60.2% and 23.1% in placebo, 31.3% and 41.8% in bebtelovimab alone arms, and 58.3% and 21.9% in the bebtelovimab with bamlanivimab and etesevimab arms, respectively. Gamma and Mu infections comprised 5.6% and 3.8% of the total infections respectively. Subjects infected with Beta, Delta [+K417N], Iota, and Lambda variants were the minority with 0.5%, 0.8%, 0.7%, and 0.5% total infections, respectively. All other subjects in the trial had SARS-CoV-2 infections from either non-WHO classified viruses (3.3%), or the lineage was not able to be determined based on the baseline sequence data (6.4%). Detection of viral variants with a 5-fold or greater reduction in susceptibility to bebtelovimab at baseline has been rare, with only one G446V substitution (8-fold shift) observed transiently out of 611 subjects in the BLAZE-4 (Arms 9-14) study that had baseline sequencing available (0.2%, 1/611).

Analysis of treatment-emergent variants focused on changes at amino acid positions with known phenotypically confirmed bebtelovimab-associated variations (i.e., K444, V445, G446, and P499) in serial viral samples obtained in the BLAZE-4 (Arms 9-14) bebtelovimab Phase 2 Study. Treatment-emergent substitutions detected at ≥15% or ≥50% allele fractions at these positions included K444E/N, V445G, G446V, and P499H/R. These substitutions resulted in a 5-fold or greater reduction in susceptibility to bebtelovimab in pseudotyped VLP assays: K444E (>862), K444N (>1,901-fold), V445G (>730-fold), G446V (8-fold), P499H (>1,606-fold), and P499R (>1,870-fold). Additional treatment-emergent substitutions detected at ≥15% or >50% allele fractions outside the epitope in at least 2 subjects included C379F (n=2) and G404C (n=2), seen in bebtelovimab in combination with bamlanivimab and etesevimab arms.

Considering all substitutions detected at ≥15% allele fraction at positions K444, V445, G446, and P499, 5.5% (11/199) of subjects treated with bebtelovimab alone harbored a variant that was treatment-emergent. This was more frequent than observed in the placebo arm (0%, 0/112), or when bebtelovimab was administered together with bamlanivimab and etesevimab (0.3%, 1/312). The appearance of these treatment-emergent bebtelovimab resistance-associated substitutions was associated with higher viral loads in the subjects in whom they were detected, but none of these subjects were hospitalized. The majority of the variants were first detected on Day 5 (n=3) and Day 7 (n=6) following treatment initiation.

It is possible that bebtelovimab resistance-associated variants could have cross-resistance to other mAbs targeting the receptor binding domain of SARS-CoV-2. The clinical impact is not known.

Immune Response Attenuation

There is a theoretical risk that antibody administration may attenuate the endogenous immune response to SARS-CoV-2 and make patients more susceptible to re-infection.

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