Science - 06.12.2019

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further examination of the biochemical proper-
ties and specificities of the naïve Abs. Twenty-
three HMP Fabs bound to N332-GT1 and/or
N332-GT2 by SPR and did not bind detect-
ably to the MD39 native-like trimer, demon-


strating proper N332 glycan supersite epitope
specificity (Fig. 3, E and F). These SPR-
validated epitope-specific clones were highly
enriched for VL3-25, VL3-1, or the closely related
VL3-10 (Fig. 3E). Thus, the protein design

strategy resulted in Envtrimersthatcould
successfully bind human naïve B cells with
BG18-like LCs.
All N332-supersite bnAbs identified to date
require a long HCDR3 [≥20 amino acids (aa)],

Steichenet al.,Science 366 , eaax4380 (2019) 6 December 2019 5of13


45

N332-GT2 AF647

N332-GT2 AF488

N332-KO AF568

N332-GT2 A488

0.059

12.4

79.0

95.8

B220+ GT2++

BG18gH

WT

Non-immunized

VH

genes

VL

genes IGKV6-32*01
IGKV4-80*01
IGKV4-79*01
IGKV4-69*01

IGKV4-68*01
IGKV4-55*01
IGKV3-4*01
IGKV3-10*01
IGKV4-61*01
IGKV3-5*01
IGKV3-1*01

IGKV12-41*01
IGKV4-57-1*01

IGKV10-96*01

IGKV12-46*01

IGKV10-94*01

IGKV12-44*01

48

48
20.8%

20.9%

12.5%

10.
5%

Frequency of B cells (%)

GT2++KO-

Day8
post-immunization

Day42
post-immunization

VH

genes

VL

genes Tree scale: 0.01

GT2 MD39

0.0

12.5

25.0

GT2 MD39

0.00

0.63

1.25

*
**

CD45.2 GT2++KO- CD45.1 GT2++KO-

Frequency of B cells [%] Frequency of B cells [%]

GC formation CD45.2

IgG ED

50

Immunogen:
GT2

GT2KOGT2KOGT2KOGT2KO

1×10-1

1×10^0

1×10^1

1×10^2

1×10^3

1×10^4

1×10^5

***

Immunogen:
MD39

0

5

10

15

BG18gH
WT

134

134

Day8
post-immunization

Day42
post-immunization

GT2-AF488 GT2-AF488

GT2-AF647 GT2-AF647

ELISA

GT2 MD39

0.00

0.40

0.80

*
**

GT2 MD39

0.00

0.06

0.12

*
ns

IGKV12-41*01
IGKV4-57-1*01

IGKV12-46*01
IGKV12-44*01

IGHV4-4*02

IGHV4-4*02

45

BC

F

A

HI

D E

G

J K
IC50
(μg/mL)
<0.01
0.01 - 0.1
0.1 - 1
1 - 50
>50

BG505_V1

mod

SF162P3_V

1 mod

AC10_V

1 mod
AD8_V1

mod

BG505_7mutBG505_MD39N332-GT2_KO

1×10^1

1×10^2

1×10^3

1×10^4

1×10^5

KD

(nM) d42_iGL
d42

naive

7m

ut

BG505_MD39N332-GT2_KO

d42_iGL
d42

naive

KD

(nM)

Ab BG505 T332N BG505-V1mod
d42.4 >50 3.3
d42.8 >50 0.06
d42.13 >50 1.61
d42.11 >50 0.45
d42.12 >50 0.1
d42.10 >50 ND
d42.iGL1 >50 >50
d42.iGL2 >50 >50
d42.iGL3 >50 >50
d42.iGL4 >50 >50
d42.iGL5 >50 >50
naive1 >50 >50
naive2 >50 >50
naive3 >50 >50
naive4 >50 >50
naive5 >50 >50
BG18 0.005 0.006

RESEARCH | RESEARCH ARTICLE


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