Key Points

  • Regnase-1 deficiency enhances CAR–T-cell persistence and CAR-T–mediated antitumor immunity in murine and human xenograft B-ALL models.

  • Regnase-1 targets Tcf7 mRNA to inhibit formation of TPEX cells critical for CAR–T-cell recall responses and survival.

Abstract

Chimeric antigen receptor (CAR)–T-cell therapeutic efficacy is associated with long-term T-cell persistence and acquisition of memory. Memory-subset formation requires T-cell factor 1 (TCF-1), a master transcription factor for which few regulators have been identified. Here, we demonstrate using an immune-competent mouse model of B-cell acute lymphoblastic leukemia (ALL; B-ALL) that Regnase-1 deficiency promotes TCF-1 expression to enhance CAR–T-cell expansion and memory-like cell formation. This leads to improved CAR-T–mediated tumor clearance, sustained remissions, and protection against secondary tumor challenge. Phenotypic, transcriptional, and epigenetic profiling identified increased tumor-dependent programming of Regnase-1–deficient CAR-T cells into TCF-1+ precursor exhausted T cells (TPEX) characterized by upregulation of both memory and exhaustion markers. Regnase-1 directly targets Tcf7 messenger RNA (mRNA); its deficiency augments TCF-1 expression leading to the formation of TPEX that support long-term CAR–T-cell persistence and function. Regnase-1 deficiency also reduces exhaustion and enhances the activity of TCF-1 CAR-T cells. We further validate these findings in human CAR-T cells, where Regnase-1 deficiency mediates enhanced tumor clearance in a xenograft B-ALL model. This is associated with increased persistence and expansion of a TCF-1+ CAR–T-cell population. Our findings demonstrate the pivotal roles of TPEX, Regnase-1, and TCF-1 in mediating CAR–T-cell persistence and recall responses, and identify Regnase-1 as a modulator of human CAR–T-cell longevity and potency that may be manipulated for improved therapeutic efficacy.

REFERENCES

1.
June
CH
,
Sadelain
M
.
Chimeric antigen receptor therapy
.
N Engl J Med
.
2018
;
379
(
1
):
64
-
73
.
2.
Maude
SL
,
Frey
N
,
Shaw
PA
, et al
.
Chimeric antigen receptor T cells for sustained remissions in leukemia
.
N Engl J Med
.
2014
;
371
(
16
):
1507
-
1517
.
3.
Xu
X
,
Sun
Q
,
Liang
X
, et al
.
Mechanisms of relapse after CD19 CAR T-cell therapy for acute lymphoblastic leukemia and its prevention and treatment strategies
.
Front Immunol
.
2019
;
10
:
2664
.
4.
Sommermeyer
D
,
Hudecek
M
,
Kosasih
PL
, et al
.
Chimeric antigen receptor-modified T cells derived from defined CD8+ and CD4+ subsets confer superior antitumor reactivity in vivo
.
Leukemia
.
2016
;
30
(
2
):
492
-
500
.
5.
Fraietta
JA
,
Lacey
SF
,
Orlando
EJ
, et al
.
Determinants of response and resistance to CD19 chimeric antigen receptor (CAR) T cell therapy of chronic lymphocytic leukemia [published correction appears in Nat Med. 2021;27(3):561]
.
Nat Med
.
2018
;
24
(
5
):
563
-
571
.
6.
McLellan
AD
,
Ali Hosseini Rad
SM
.
Chimeric antigen receptor T cell persistence and memory cell formation
.
Immunol Cell Biol
.
2019
;
97
(
7
):
664
-
674
.
7.
Busch
DH
,
Fräßle
SP
,
Sommermeyer
D
,
Buchholz
VR
,
Riddell
SR
.
Role of memory T cell subsets for adoptive immunotherapy
.
Semin Immunol
.
2016
;
28
(
1
):
28
-
34
.
8.
Long
AH
,
Haso
WM
,
Shern
JF
, et al
.
4-1BB costimulation ameliorates T cell exhaustion induced by tonic signaling of chimeric antigen receptors
.
Nat Med
.
2015
;
21
(
6
):
581
-
590
.
9.
Zebley
CC
,
Gottschalk
S
,
Youngblood
B
.
Rewriting history: epigenetic reprogramming of CD8+ T cell differentiation to enhance immunotherapy
.
Trends Immunol
.
2020
;
41
(
8
):
665
-
675
.
10.
Xu
Y
,
Zhang
M
,
Ramos
CA
, et al
.
Closely related T-memory stem cells correlate with in vivo expansion of CAR.CD19-T cells and are preserved by IL-7 and IL-15
.
Blood
.
2014
;
123
(
24
):
3750
-
3759
.
11.
Gardner
RA
,
Finney
O
,
Annesley
C
, et al
.
Intent-to-treat leukemia remission by CD19 CAR T cells of defined formulation and dose in children and young adults
.
Blood
.
2017
;
129
(
25
):
3322
-
3331
.
12.
Yang
S
,
Ji
Y
,
Gattinoni
L
, et al
.
Modulating the differentiation status of ex vivo-cultured anti-tumor T cells using cytokine cocktails
.
Cancer Immunol Immunother
.
2013
;
62
(
4
):
727
-
736
.
13.
Fraietta
JA
,
Nobles
CL
,
Sammons
MA
, et al
.
Disruption of TET2 promotes the therapeutic efficacy of CD19-targeted T cells
.
Nature
.
2018
;
558
(
7709
):
307
-
312
.
14.
Feucht
J
,
Sun
J
,
Eyquem
J
, et al
.
Calibration of CAR activation potential directs alternative T cell fates and therapeutic potency
.
Nat Med
.
2019
;
25
(
1
):
82
-
88
.
15.
Zheng
W
,
O’Hear
CE
,
Alli
R
, et al
.
PI3K orchestration of the in vivo persistence of chimeric antigen receptor-modified T cells
.
Leukemia
.
2018
;
32
(
5
):
1157
-
1167
.
16.
Lynn
RC
,
Weber
EW
,
Sotillo
E
, et al
.
c-Jun overexpression in CAR T cells induces exhaustion resistance
.
Nature
.
2019
;
576
(
7786
):
293
-
300
.
17.
Eyquem
J
,
Mansilla-Soto
J
,
Giavridis
T
, et al
.
Targeting a CAR to the TRAC locus with CRISPR/Cas9 enhances tumour rejection
.
Nature
.
2017
;
543
(
7643
):
113
-
117
.
18.
Mino
T
,
Murakawa
Y
,
Fukao
A
, et al
.
Regnase-1 and roquin regulate a common element in inflammatory mRNAs by spatiotemporally distinct mechanisms
.
Cell
.
2015
;
161
(
5
):
1058
-
1073
.
19.
Uehata
T
,
Iwasaki
H
,
Vandenbon
A
, et al
.
Malt1-induced cleavage of regnase-1 in CD4(+) helper T cells regulates immune activation
.
Cell
.
2013
;
153
(
5
):
1036
-
1049
.
20.
Matsushita
K
,
Takeuchi
O
,
Standley
DM
, et al
.
Zc3h12a is an RNase essential for controlling immune responses by regulating mRNA decay
.
Nature
.
2009
;
458
(
7242
):
1185
-
1190
.
21.
Mao
R
,
Yang
R
,
Chen
X
,
Harhaj
EW
,
Wang
X
,
Fan
Y
.
Regnase-1, a rapid response ribonuclease regulating inflammation and stress responses
.
Cell Mol Immunol
.
2017
;
14
(
5
):
412
-
422
.
22.
Kurachi
M
,
Barnitz
RA
,
Yosef
N
, et al
.
The transcription factor BATF operates as an essential differentiation checkpoint in early effector CD8+ T cells
.
Nat Immunol
.
2014
;
15
(
4
):
373
-
383
.
23.
Wei
J
,
Long
L
,
Zheng
W
, et al
.
Targeting REGNASE-1 programs long-lived effector T cells for cancer therapy
.
Nature
.
2019
;
576
(
7787
):
471
-
476
.
24.
He
R
,
Hou
S
,
Liu
C
, et al
.
Follicular CXCR5- expressing CD8(+) T cells curtail chronic viral infection [published correction appears in Nature. 2016;540(7633):470]
.
Nature
.
2016
;
537
(
7620
):
412
-
428
.
25.
Miller
BC
,
Sen
DR
,
Al Abosy
R
, et al
.
Subsets of exhausted CD8+ T cells differentially mediate tumor control and respond to checkpoint blockade [published correction appears in Nat Immunol. 2019;20(11):1556]
.
Nat Immunol
.
2019
;
20
(
3
):
326
-
336
.
26.
Chen
Z
,
Ji
Z
,
Ngiow
SF
, et al
.
TCF-1-centered transcriptional network drives an effector versus exhausted CD8 T cell-fate decision
.
Immunity
.
2019
;
51
(
5
):
840
-
855.e5
.
27.
Alfei
F
,
Kanev
K
,
Hofmann
M
, et al
.
TOX reinforces the phenotype and longevity of exhausted T cells in chronic viral infection
.
Nature
.
2019
;
571
(
7764
):
265
-
269
.
28.
Scott
AC
,
Dündar
F
,
Zumbo
P
, et al
.
TOX is a critical regulator of tumour-specific T cell differentiation
.
Nature
.
2019
;
571
(
7764
):
270
-
274
.
29.
Yao
C
,
Sun
HW
,
Lacey
NE
, et al
.
Single-cell RNA-seq reveals TOX as a key regulator of CD8+ T cell persistence in chronic infection
.
Nat Immunol
.
2019
;
20
(
7
):
890
-
901
.
30.
Blank
CU
,
Haining
WN
,
Held
W
, et al
.
Defining “T cell exhaustion”
.
Nat Rev Immunol
.
2019
;
19
(
11
):
665
-
674
.
31.
Kallies
A
,
Zehn
D
,
Utzschneider
DT
.
Precursor exhausted T cells: key to successful immunotherapy?
Nat Rev Immunol
.
2020
;
20
(
2
):
128
-
136
.
32.
Utzschneider
DT
,
Charmoy
M
,
Chennupati
V
, et al
.
T cell factor 1-expressing memory-like CD8(+) T cells sustain the immune response to chronic viral infections
.
Immunity
.
2016
;
45
(
2
):
415
-
427
.
33.
Zhou
X
,
Yu
S
,
Zhao
DM
,
Harty
JT
,
Badovinac
VP
,
Xue
HH
.
Differentiation and persistence of memory CD8(+) T cells depend on T cell factor 1
.
Immunity
.
2010
;
33
(
2
):
229
-
240
.
34.
Raghu
D
,
Xue
HH
,
Mielke
LA
.
Control of lymphocyte fate, infection, and tumor immunity by TCF-1
.
Trends Immunol
.
2019
;
40
(
12
):
1149
-
1162
.
35.
Platt
RJ
,
Chen
S
,
Zhou
Y
, et al
.
CRISPR-Cas9 knockin mice for genome editing and cancer modeling
.
Cell
.
2014
;
159
(
2
):
440
-
455
.
36.
Churchman
ML
,
Evans
K
,
Richmond
J
, et al
.
Synergism of FAK and tyrosine kinase inhibition in Ph+ B-ALL
.
JCI Insight
.
2016
;
1
(
4
):
e86082
.
37.
Tan
H
,
Yang
K
,
Li
Y
, et al
.
Integrative proteomics and phosphoproteomics profiling reveals dynamic signaling networks and bioenergetics pathways underlying T cell activation
.
Immunity
.
2017
;
46
(
3
):
488
-
503
.
38.
Abdelsamed
HA
,
Moustaki
A
,
Fan
Y
, et al
.
Human memory CD8 T cell effector potential is epigenetically preserved during in vivo homeostasis
.
J Exp Med
.
2017
;
214
(
6
):
1593
-
1606
.
39.
Imai
C
,
Mihara
K
,
Andreansky
M
, et al
.
Chimeric receptors with 4-1BB signaling capacity provoke potent cytotoxicity against acute lymphoblastic leukemia
.
Leukemia
.
2004
;
18
(
4
):
676
-
684
.
40.
Kalos
M
,
Levine
BL
,
Porter
DL
, et al
.
T cells with chimeric antigen receptors have potent antitumor effects and can establish memory in patients with advanced leukemia
.
Sci Transl Med
.
2011
;
3
(
95
):
95ra73
.
41.
Jansen
CS
,
Prokhnevska
N
,
Master
VA
, et al
.
An intra-tumoral niche maintains and differentiates stem-like CD8 T cells
.
Nature
.
2019
;
576
(
7787
):
465
-
470
.
42.
Kurtulus
S
,
Madi
A
,
Escobar
G
, et al
.
Checkpoint blockade immunotherapy induces dynamic changes in PD-1-CD8+ tumor-infiltrating T cells
.
Immunity
.
2019
;
50
(
1
):
181
-
194.e6
.
43.
Utzschneider
DT
,
Legat
A
,
Fuertes Marraco
SA
, et al
.
T cells maintain an exhausted phenotype after antigen withdrawal and population reexpansion
.
Nat Immunol
.
2013
;
14
(
6
):
603
-
610
.
44.
Siddiqui
I
,
Schaeuble
K
,
Chennupati
V
, et al
.
Intratumoral Tcf1+PD-1+CD8+ T cells with stem-like properties promote tumor control in response to vaccination and checkpoint blockade immunotherapy
.
Immunity
.
2019
;
50
(
1
):
195
-
211.e10
.
45.
Abdelsamed
HA
,
Zebley
CC
,
Nguyen
H
, et al
.
Beta cell-specific CD8+ T cells maintain stem cell memory-associated epigenetic programs during type 1 diabetes
.
Nat Immunol
.
2020
;
21
(
5
):
578
-
587
.
46.
McLane
LM
,
Abdel-Hakeem
MS
,
Wherry
EJ
.
CD8 T cell exhaustion during chronic viral infection and cancer
.
Annu Rev Immunol
.
2019
;
37
:
457
-
495
.
47.
Kratchmarov
R
,
Magun
AM
,
Reiner
SL
.
TCF1 expression marks self-renewing human CD8+ T cells
.
Blood Adv
.
2018
;
2
(
14
):
1685
-
1690
.
48.
Jeannet
G
,
Boudousquié
C
,
Gardiol
N
,
Kang
J
,
Huelsken
J
,
Held
W
.
Essential role of the Wnt pathway effector Tcf-1 for the establishment of functional CD8 T cell memory
.
Proc Natl Acad Sci USA
.
2010
;
107
(
21
):
9777
-
9782
.
49.
Utzschneider
DT
,
Gabriel
SS
,
Chisanga
D
, et al
.
Early precursor T cells establish and propagate T cell exhaustion in chronic infection
.
Nat Immunol
.
2020
;
21
(
10
):
1256
-
1266
.
50.
Yu
D
,
Ye
L
.
A portrait of CXCR5+ follicular cytotoxic CD8+ T cells
.
Trends Immunol
.
2018
;
39
(
12
):
965
-
979
.
51.
Wu
T
,
Ji
Y
,
Moseman
EA
, et al
.
The TCF1-Bcl6 axis counteracts type I interferon to repress exhaustion and maintain T cell stemness
.
Sci Immunol
.
2016
;
1
(
6
):
eaai8593
.
52.
Beltra
JC
,
Manne
S
,
Abdel-Hakeem
MS
, et al
.
Developmental relationships of four exhausted CD8+ T cell subsets reveals underlying transcriptional and epigenetic landscape control mechanisms
.
Immunity
.
2020
;
52
(
5
):
825
-
841.e8
.
53.
Sade-Feldman
M
,
Yizhak
K
,
Bjorgaard
SL
, et al
.
Defining T cell states associated with response to checkpoint immunotherapy in melanoma [published correction appears in Cell. 2019;176(1-2):404]
.
Cell
.
2018
;
175
(
4
):
998
-
1013.e20
.
54.
Godec
J
,
Tan
Y
,
Liberzon
A
, et al
.
Compendium of immune signatures identifies conserved and species-specific biology in response to inflammation
.
Immunity
.
2016
;
44
(
1
):
194
-
206
.
You do not currently have access to this content.

Sign in via your Institution

Sign In