41591_2022_1762_MOESM1

Safety and tolerability of AAV8 delivery of a

broadly neutralizing antibody in adults

living with HIV: a phase 1, dose-escalation

trial

In the format provided by the

authors and unedited

Supplementary information

https://doi.org/10.1038/s41591-022-01762-x

Supplemental Tables and Figures.

Supplemental Table 1. Summary of reasons for ineligibility

Cause

Number of Candidates Found

Ineligible

Hypertension

AAV8 Ab positive*

Active drug use

Elevated creatinine

Weight > 115 kg

Inadequate venous access

Renal disease

Cardiovascular disease

VL > 50 copies/mL

Significant chronic pain

Chronic hepatitis

Unstable ARV regimen

Mental health concerns

Unable to provide ID

Previous receipt of mAb

CD4+ < 300

Stargdardt's disease

Epilepsy

Autoimmune condition

Neurosyphilis

Elevated ALT

Chronic respiratory condition

*Criteria for AAV8 seropositivity was modified by protocol amendment because of NHP data suggesting

that low level pre-existing AAV8 seropositivity did not affect transduction. Based on these data the final

exclusion criteria for pre-existing seropositivity was increased to a titer >1:90. Data presented here

includes disqualification of volunteers for values of <1:90.

Supplemental

Table 2

. Demographic characteristics of study participants

Category

Subcategory

Group 1

(N=3)

Group 2

(N=2)

Group 3

(N=3)

Overall

(N=8)

N (%)

Gender

Male

Female

3 (100)

0 (0)

2 (100)

0 (0)

1 (33)

2 (66)

6 (75)

2 (25)

Race

Asian

Black/African

White

0 (0)

1 (33)

2 (66.7)

0 (0)

1 (50)

0 (0)

3 (100)

0 (0)

5 (62.5)

3 (37.5)

Ethnicity

Non

Hispanic/Latino

3(100)

0(0)

2(100)

0(0)

3(100)

0(0)

8(100)

0(0)

Median Age

Median Years

(range)

56 (36

60)

41 (30

52)

52 (32

56)

52 (30

60)

Median Weight

Median Kg

weight (range)

79 (66

82)

74 (73

75)

82 (71

91)

77 (66

91)

Supplemental Table 3. Clinical

characteristic of participants at enrollment

Participant

CD4 (cells/μl)

VL (copies/ml)

Antiretroviral Therapy

Participant A

523

Tenofovir Alafenamide

Emtricitabine

Elvitegravir/Cobicistat

Participant B

446

<20

Abacavir

Efavirenz

Fosamprenavir/Ritonavir

Participant C

532

<20

Tenofovir Alafenamide

Emtricitabine

Rilpivirine

Participant D

950

<20

Abacavir

Lamivudine

Dolutegravir

Participant E

351

<20

Tenofovir Alafenamide

Emtricitabine

Darunavir/Ritonavir

Participant F

641

<20

Tenofovir Disoproxil

Emtricitabine

Rilpivirine

Participant G

407

<20

Tenofovir Alafenamide

Emtricitabine

Darunavir/Ritonavir

Participant H

797

<20

Abacavir

Lamivudine

Dolutegravir

Supplemental Table 4. Participant IgG1 Allotype and

Association with ADA

Participant

GM1

GM3/17

ADA reponse

Participant A*

Participant B

Participant C

Participant D

Participant E*

Participant F

Participant G

Participant H*

-/-

+/+

-/-

+/+

3+/3+

17+/17+

3+/3+

17+/17+

Tier 1/2+ Tier 3

Tier 1/2- Tier 3-

Tier 1/2+ Tier 3-

Tier 1/2- Tier 3-

Tier 1/2+ Tier 3-

*Indicates Allotype from a participant with a positive VRC07 ADA

response

Supplemental Table 5.

Geometric means

2 SD for

ex vivo

and

in vivo

produced VRC07

Pseudovirus

Geometric mean

2 SD

+ 2 SD

ID50s for

ex vivo

produce VRC07 (μg/ml)

001428

0.0093

0.0032

0.0271

JRFLJB

0.0147

0.0038

0.0570

Q842

0.0304

0.0147

0.0632

T33

0.0146

0.0086

0.0247

TZBD

0.0355

0.0194

0.0647

ID50s from purified IgGs from all

in vivo

produced VRC07 (μg/ml)

001428

0.0081

0.0018

0.0364

JRFLJB

0.0128

0.0026

0.0641

Q842

0.0193

0.0026

0.144

T33

0.0375

0.027

0.0521

TZBD

0.0357

0.0178

0.0711

Supplemental Figure 1 AAV8 capsid serotiters for 48 pre-enrollment trial candidates.

Titers are

plotted as inverse serum dilution

Supplemental Figure. 2 Longitudinal serum AST and ALT values after IM injection of 5 x 10

, 5 x 10

and 2.5 x 10

AAV8-VRC07 vg/kg.

Measured ALT is shown in column a), measured AST is shown in

column b). Data from individual participants are as shown in the figure legend. Participants A-C

received 5 x 10

, Participants D and E received 5 x 10

, and participants F-H received 2.5 x 10

AAV8-

VRC07 vg/kg. Limits of normal values for a) AST and b) ALT in males are shown in shading.

Supplemental Figure 3a Frequency (%) of CD4

and CD8

T cells producing IFNγ after stimulation with

15mer peptides overlapping by 11 amino acids covering capsid protein VP1, VP2 and VP3.

Activity was

measured using intracellular cytokine staining of cells incubated for 6h in the presence of monensin and

brefeldin A. IFNγ responses for individual subjects are as indicated in the figure legend. Individual

participants are as indicated in the figure legend. b) Gating strategy for the quantificaion of Intracellular

IFNγ staining in CD4 and CD8 T cells in response to stimulation by an AAV8 peptide pool containing

15mer overlapping by 11 amino acids covering VP1, VP2, and VP3.

Supplemental Fig. 4 Longitudinal inverse anti-AAV8 capsid serotiters before and between 2 -104

weeks after IM injection of 5 x 10

, 5 x 10

and 2.5 x 10

AAV8-VRC07 vg/kg.

Individual participant

values are as shown in the figure legend. Participants A-C received 5 x 10

, participants D and E receive

5 x 10

and participants F-H received 2.5 x 10

vg AAV8-VRC07/kg.

Supplemental Figure 5 Longitudinal HIV viral loads for participants in the 5 x 10

, 5 x 10

and 2.5 x

AAV8-VRC07 vg/kg dose groups.

Participants A-C received 5 x 10

, participants D and E received

5 x 10

and participant F-H received 2.5 x 10

AAV8-VRC07 vg/kg. Data from individual participants are

as indicated in the figure legend. Lower level of quantitation was 20 copies of HIV RNA/ml.

Supplemental Figure 6 Longitudinal CD4 T cell count for participants in 5 x 10

, 5 x 10

and 2.5 x 10

AAV8-VRC07 vg /kg dose groups.

Participants A-C received 5 x 10

, participants D and E received 5 x

and participants F-H received 2.5 x 10

AAV8-VRC07 vg/kg. Individual participants are as shown in

the figure legend.

Supplemental Figure 7 Longitudinal tier 1 VRC07 ADA (left Y axis) and Fab directed ADA (right Y axis)

activity.

ADA activity is shown as measured electrochemiluminescence (ECL) in 1:30 dilutions of serum

from participants in the 5 x 10

, 5 x 10

and 2.5 x 10

AAV8-VRC07 vg/kg dose groups. Tier 1 ADA

responses are shown on the left y axis, tier 1 ADA responses directed against the Fab fragment are

shown on the right y axis.

Supplemental Figure 8 Longitudinal tier 2 pseudovirus neutralization by purified IgG containing

VRC07.

Percent neutralization of tier 2 pseudovirus in participant A-G, by purified IgG containing VRC07,

is shown on the left axis. Measured VRC07 contained in each assay is shown on the x axis.

Neutralization curves for each purified IgG sampled at the indicated timepoints are overlaid with the

neutralization curve of

in vitro

-purified VRC07 (black lines). Pseudovirus neutralized is shown in the left

margin. Data points for neutralization assays of ex vivo produced VRC07 represent the average of 6

replicate assays, error bars are + SD. Data points for

in vivo

produced VRC07 represent a single assay.

Supplemental Fig. 9 Comparison of longitudinal pseudoviral IC50s for

in vivo

produced and

in vitro

produced and purified VRC07.

Longitudinal IC50s determined for

in vivo

produced VRC07 from purified

IgGs from seven different participants are as shown in the figure legend to the right of the graph as

compared to IC50s determined for

ex vitro

produced VRC07 shown in black dots shown at the left of

each segment. Shaded areas show +2 SD around the geometric mean of the

ex vitro

samples. In total

IgG was purified from 120 different plasma sample. From those samples 77 samples contained adequate

VRC07 to determine the IC50s for pseudoviral neutralizations studies. Determinations of

ex vivo

produced and

in vitro

produced VRC07 IC50s were performed using the same pseudovirus lots. Each

data point shown represents one assay.