nature medicine
https://doi.org/10.1038/s41591-024-02979-8
Artic�e
PSCA-CAR T cell therapy in metastatic
castration-resistant prostate cancer: a phase
1 trial
In the format provided by the
authors and unedited
Supp�ementary information
Supplementary Figure 1
.
PSCA
-
CAR construct.
Humanized
A11
scFv
△
CH2
4
-
1BB
CD3z
CD4 TM
Supplementary
Figure
2
. Peripheral blood CAR T cell expansion and phenotype. (a)
Representative patient per DL showing peripheral blood CAR T cell persistence following T cell
infusion.
(b
-
d)
Expression of PD1 (b), CD25 (c), and CD95 (d) among CD8+CAR+ T cells in
peripheral blood at peak CAR T cell expansion for each patient in each DL
(DL1 n = 3, DL2 n =
6, DL3 n = 5)
.
Data are presented as mean values ±
SEM
.
DL1
DL2
DL3
Wk
0
Wk
1
Wk
2
Wk
4
Pre
-
LD
1.7%
3.1%
6.4%
1.9%
0.3%
1.5%
1.6%
5.1%
5.1%
3.2%
1.2%
0.2%
0.9%
0.3%
1.3%
0.5%
10
0
10
1
10
2
10
3
10
4
10
5
10
0
10
1
10
2
10
3
10
4
10
5
CD3
-
VioGreen
CD19
-
PECy7
Wk
3
b
d
c
DL1
DL2
DL3
0
10
20
30
CD8+CAR+PD1+ cells
DL1
DL2
DL3
0
20
40
60
80
100
CD8+CAR+CD25+ cells
DL1
DL2
DL3
0
20
40
60
80
100
CD8+CAR+CD95+ cells
a
Supplementary
Figure
3
. Peripheral blood CTC analysis in UPN388.
0
20
40
60
80
Peripheral blood
(cells/mL)
D/CK/CD/V
D/CK
D
D/CD
D/CD/V
D/V
D/CK/CD
D/CK/V
Before
infusion
1 month
after
infusion
CK+ cells/mL:
23
12
Supplementary
Figure
4
. Flow cytometric analysis of peripheral blood CD8+PD1+ CAR and
non
-
CAR T
cells in UPN388. (a)
Flow cytometric analysis of expression of CD3+CD19+ CAR
and CD3+CD19
-
non
-
CAR T cells in peripheral blood for UPN 388.
(b
-
c)
Expression of PD1
among CD3+CAR
-
non
-
CAR (b) and CD3+CAR+ CAR (c) T cells.
Iso
Day 0
Day 7
Day 14
Day 21
Day 224
Day 28
0%
100%
0%
100%
3%
97%
35%
65%
9%
91%
1%
99%
4%
96%
0%
0%
0%
1.7%
9.5%
16.0%
9.3%
0%
0%
0%
3.6%
15
.3
%
36.6%
31.5%
10
1
10
2
10
3
10
4
10
5
10
0
10
1
10
2
10
3
10
4
CD3
-
VioGreen
CD19
-
PECy7
10
5
10
2
10
3
10
4
10
5
10
0
10
1
10
2
10
3
10
4
CD8
-
APCCy7
PD1
-
FITC
10
5
10
1
10
2
10
3
10
4
10
5
10
0
10
1
10
2
10
3
10
4
CD8
-
APCCy7
PD1
-
FITC
10
5
10
1
Gated on CD3+
Gated on CD3+CAR
-
Gated on CD3+CAR+
a
b
c
Supplementary Table
1
. CTC analysis in bone marrow.
DL1 (N=3)
DL2 (N=5)
DL3 (N=4*)
Mean +/
-
STD
Total CK+
5.43 +/
-
7.47
-
12.33
+/
-
19.85
-
13.80
+/
-
77.09
D|CK|V|CD
2.51 +/
-
4.71
-
6.39
+/
-
19.15
-
12.70
+/
-
30.38
D|CK|CD
3.61
+/
-
1.58
-
0.81
+/
-
2.90
2.04
+/
-
45.52
D|CK
-
0.83
+/
-
3.08
-
5.81
+/
-
11.67
-
3.21
+/
-
5.92
D|CK|V
0.15
+/
-
0.82
0.68
+/
-
1.16
0.07
+/
-
0.53
Median
Total CK+
1.20
-
3.34
-
11.15
D|CK|V|CD
-
0.82
-
1.04
-
1.72
D|CK|CD
3.85
-
1.40
-
3.56
D|CK
-
0.80
-
0.99
-
1.82
D|CK|V
0.00
0.00
0.00
*The
patient
that
received
2
cycles
of
therapy
is
treated
as
2
unique
timepoints
.
Supplementary Table 2. Dose
-
limiting toxicities (DLTs) and cytokine release syndrome
(CRS).
Cohort
N
DLTs
CRS
DL1: 100M
3
0 (0%)
95%CI (0%,71%)
1 Grade 2
DL2:
LD + 100M
6
2 (33%)
95%CI (4%,78%)
1 Grade 1
1 Grade 2
DL3:
Modified LD + 100M
5
0 (0%)
95%CI (0%,52%)
2 Grade 1
Supplementary Table
3
. MACSQuant 10 antibody list.
MACSQuant
10
antibody
list
Marker
Fluorophore
Clone
Manufacturer
Catalog #
PD1
FITC
EH12.2H7
BioLegend
329936
CD25
APC
2A3
BD Biosciences
340939
CD137
PE
4B4-1
BD Biosciences
555956
CD19
PE-Cy7
SJ25C1
BD Biosciences
557835
CD8
APC-Cy7
SK1
BD Biosciences
348793
CD3
BV510
UCHT1
BD Biosciences
563109
CD95
PerCP-Cy5.5
DX2
BD Biosciences
561655
CD33
FITC
P67.6
BD Biosciences
340533
CD11b
APC
M1/70.15.11.5
Miltenyi Biotec Inc.
130-113-231
CD86
PE
2331 (FUN-1)
BD Pharmingen
555658
CD14
PE-Cy7
MφP9
BD Biosciences
562698
HLA-DR
APC-eFluor 780
LN3
eBioscience
47-9956-42
CD16
BV510
B73.1
BD Horizon
740203
CD15
PE-Cy5
W6D3
BioLegend
323014
Supplementary Table
4
. Cytek
Aurora 3 antibody list.
Cytek
Aurora
3
antibody
list
Marker
Fluorophore
Clone
Manufacturer
Catalog #
CD3
Alexa Fluor 532
UCHT1
eBioscience
58003842
CD4
PerCP
SK3
BD Biosciences
347324
CD8
BV570
RPA-T8
BioLegend
301038
CD45RA
BV650
HI100
BD Biosciences
563963
CD45
Alexa Fluor 700
HI30
BD Biosciences
560566
CD62L
BV711
SK11
BD Biosciences
565040
CD19
V450
HIB19
BD Biosciences
560353
CD39
BV510
A1
BioLegend
328220
CD69
BV750
FN50
BioLegend
310954
CD14
FITC
MphiP9
BioLegend
363508
CX3CR1
PE-Cy7
2A9-1
BioLegend
341612
LD
7AAD
-
eBioscience
00-6993-50
CXCR3
PE
G025H7
BioLegend
353706
PD-1
BV421
MIH4
BD Biosciences
BDB564323
CD28
APC-Cy7
CD28.2
BioLegend
302966
LAG-3
APC
3DS223H
eBioscience
17-2239-42
Tim3
PE-Cy5.5
F38-2E2
eBioscience
35-3109-42
CD16
BV786
3G8
BD Biosciences
563689
CCR4
Alexa Flour 647
L291H4
BioLegend
359404
CD27
PE-CF594
M-T271
BD Biosciences
BDB562297
CCR7
BV605
4B12
BioLegend
353224
CD45RO
BV480
UCHL1
BD Biosciences
BDB566143
CD25
PerCPeFluor710
BC96
eBioscience
46-0259-42
PD-L1
PE-Fire810
29E.2A3
BioLegend
329755
CD95
APC-Fire 810
DX2
BioLegend
305663