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ACKNOWLEDGMENTS
We thank the University of Manchester Mass Spectrometry Service
Centre (G. Smith and E. Enston) for MALDI-TOF and high-
resolution mass spectrometry, the Swedish National Infrastructure
for Computing at the National Supercomputer Centre of Linköping
University and Stanford University and the Stanford Research
Computing Center for computational resources, and
J.-F. Lemonnier and S. Borsley for useful discussions.Funding:
This work was supported by the Engineering and Physical Sciences
Research Council (EPSRC grant EP/P027067/1); the European
Research Council (ERC advanced grant 786630); Marie
Skłodowska-Curie Actions of the European Union (individual
postdoctoral fellowship EC 746993 to F.S.); the German Research
Foundation (DFG individual postdoctoral fellowship to E.K.); the
Knut and Alice Wallenberg Foundation (individual postdoctoral
fellowship 2019.0586 to J.H.S.); and the US Department of Energy,Office of Science, Office of Basic Energy Sciences, Chemical Sciences,
Geosciences, and Biosciences Division, Catalysis Science Program to
the SUNCAT Center for Interface Science and Catalysis (F.A.-P.). D.A.L.
is a Royal Society research professor.Author contributions:Z.A., E.K.,
L.P., and F.S. planned and completed the synthetic work. J.H.S.
and F.A.-P. performed computational investigations. D.A.L. directed the
research. Z.A., E.K., L.P., F.S., J.H.S., F.A.-P., and D.A.L. analyzed the
results and wrote the manuscript.Competing interests:The authors
declare no competing interests.Data and materials availability:All
data are available in the main text or the supplementary materials.
SUPPLEMENTARY MATERIALS
science.org/doi/10.1126/science.abm9247
Supplementary Text
Figs. S1 to S128
Tables S1 to S3
References ( 56 Ð 81 )
21 October 2021; accepted 1 February 2022
10.1126/science.abm9247CORONAVIRUS
Broad antiÐSARS-CoV-2 antibody immunity induced
by heterologous ChAdOx1/mRNA-1273 vaccination
Chengzi I. Kaku1,2, Elizabeth R. Champney^1 , Johan Normark^3 , Marina Garcia^4 , Carl E. Johnson^1 ,
Clas Ahlm^3 , Wanda Christ^4 , Mrunal Sakharkar^1 , Margaret E. Ackerman2,5, Jonas Klingström^4 ,
Mattias N. E. Forsell^3 , Laura M. Walker1,6*
Heterologous prime-boost immunization strategies have the potential to augment COVID-19 vaccine efficacy.
We longitudinally profiled severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike (S)Ðspecific
serological and memory B cell (MBC) responses in individuals who received either homologous (ChAdOx1:
ChAdOx1) or heterologous (ChAdOx1:mRNA-1273) prime-boost vaccination. Heterologous messenger RNA
(mRNA) booster immunization induced higher serum neutralizing antibody and MBC responses against
SARS-CoV-2 variants of concern (VOCs) compared with that of homologous ChAdOx1 boosting. Specificity
mapping of circulating B cells revealed that mRNA-1273 boost immunofocused ChAdOx1-primed responses
onto epitopes expressed on prefusion-stabilized S. Monoclonal antibodies isolated from mRNA-1273Ðboosted
participants displayed overall higher binding affinities and increased breadth of reactivity against VOCs relative
to those isolated from ChAdOx1-boosted individuals. Overall, the results provide molecular insight into the
enhanced quality of the B cell response induced after heterologous mRNA booster vaccination.
M
ultiple safe and effective COVID-19
vaccines have been developed at an
unprecedented scale and speed. How-
ever, waning vaccine-induced immu-
nity and the emergence of severe
acute respiratory syndrome coronavirus 2
(SARS-CoV-2) variants of concern (VOCs) with
increased neutralization resistance have lim-
ited the effectiveness of currently available
vaccines ( 1 – 3 ). An early challenge in vaccine
distribution was a halt in ChAdOx1 nCoV-19/
AZD1222 (hereafter referred to as ChAdOx1)
booster immunization in certain age groups
driven by evidence of rare but serious throm-
botic events ( 4 , 5 ). To fully immunize individ-
uals who received a single dose of ChAdOx1
but were not eligible for a ChAdOx1 boost,
several countries recommended a switch to
heterologous booster vaccination with mRNA-
1273 (Moderna) or BNT162b2 (Pfizer/BioNTech).
This provided an opportunity to learn that
heterologous ChAdOx1/mRNA prime-boost im-
munization induces higher serum neutralizing
antibody titers and confers increased levels
of protection relative to homologous ChAdOx1
dosing, but the molecular basis for this differ-
ence in immunogenicity remains unknown ( 6 – 9 ).
All currently available COVID-19 vaccines are
based on the SARS-CoV-2 spike (S) protein,
which plays a key role in viral entry and is the
primary target for neutralizing antibodies.
TheSproteinexistsonthesurfaceofthe
virion in a metastable prefusion conformation,
and binding of the receptor binding domain
(RBD) to angiotensin-converting enzyme 2
(ACE2) triggers shedding of the S1 subunit
and transition of the S2 subunit to a highly
stable postfusion conformation ( 10 ). Early struc-tural and biochemical studies demonstrated
that stabilization of the spike ectodomain
through two consecutive proline substitutions
in the S2 subunit (S-2P) prevents the transi-
tion from the prefusion to postfusion state and
leads to enhanced immunogenicity in animal
models ( 11 , 12 ). Several COVID-19 vaccines en-
code S-2P, so that the protein maintains the
prefusion conformation and avoids S1 shed-
ding (for example, mRNA-1273, BNT162b2, and
Ad26.COV2.S), whereas others express wild-
type (WT) S (for example, ChAdOx1, Sputnik V,
and CoronaVac), which likely leads to expres-
sion of both pre- and postfusion conforma-
tions of S. We comprehensively interrogated
serological and circulating B cell responses
induced by prime immunization with an adeno-
virus vector–based vaccine encoding WT S
(ChAdOx1) and after a second dose of either
ChAdOx1oranmRNAvaccineexpressingS-2P
(mRNA-1273).
We recruited 55 health care workers who re-
ceived either homologous ChAdOx1:ChAdOx1
or heterologous ChAdOx1:mRNA-1273 prime-
boost vaccination for blood donation (table
S1). None of the volunteers had a documented
history of prior SARS-CoV-2 infection. Par-
ticipants received one dose of ChAdOx1 and,
9 to 12 weeks later, a second dose of either
ChAdOx1 (n= 28 participants) or mRNA-1273
(n= 27 participants). We collected the first
blood sample on the day of booster immuni-
zation to analyze ChAdOx1-primed immune
responses and a second sample 7 to 10 days
after the second dose to study the early sec-
ondary B cell response induced by homologous
or heterologous booster vaccination (Fig. 1A).
We first evaluated serum immunoglobulin
G(IgG)bindingactivityatbothsamplingtime
points. All participants mounted weak but de-
tectable SARS-CoV-2 S-specific serum IgG bind-
ingresponsesafterthefirstdoseofChAdOx1,
and homologous booster vaccination resulted
in a small but significant (4.6-fold) increase in
serum IgG binding antibodies (Fig. 1B andSCIENCEscience.org 4 MARCH 2022¥VOL 375 ISSUE 6584 1041
(^1) Adimab, Lebanon, NH 03766, USA. (^2) Thayer School of
Engineering, Dartmouth College, Hanover, NH 03755, USA.
(^3) Division of Immunology, Department of Clinical
Microbiology, Umeå University, Umeå, Sweden.^4 Centre for
Infectious Medicine, Department of Medicine Huddinge,
Karolinska Institutet, Stockholm, Sweden.^5 Geisel School of
Medicine, Dartmouth College, Hanover, NH 03755, USA.
(^6) Adagio Therapeutics, Waltham, MA 02451, USA.
*Corresponding author. Email: [email protected]
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