CAR-T cells and TRUCKs that recognize an EBNA-3C-derived epitope presented on HLA-B*35 control Epstein-Barr virus-associated lymphoproliferation.
CD8-positive t-lymphocytes
cell engineering
chimeric antigen
immunotherapy
receptors
transplantation immunology
Journal
Journal for immunotherapy of cancer
ISSN: 2051-1426
Titre abrégé: J Immunother Cancer
Pays: England
ID NLM: 101620585
Informations de publication
Date de publication:
10 2020
10 2020
Historique:
accepted:
18
09
2020
entrez:
31
10
2020
pubmed:
1
11
2020
medline:
6
10
2021
Statut:
ppublish
Résumé
Immunosuppressive therapy or T-cell depletion in transplant patients can cause uncontrolled growth of Epstein-Barr virus (EBV)-infected B cells resulting in post-transplant lymphoproliferative disease (PTLD). Current treatment options do not distinguish between healthy and malignant B cells and are thereby often limited by severe side effects in the already immunocompromised patients. To specifically target EBV-infected B cells, we developed a novel peptide-selective chimeric antigen receptor (CAR) based on the monoclonal antibody TÜ165 which recognizes an Epstein-Barr nuclear antigen (EBNA)-3C-derived peptide in HLA-B*35 context in a T-cell receptor (TCR)-like manner. In order to attract additional immune cells to proximity of PTLD cells, based on the TÜ165 CAR, we moreover generated T cells redirected for universal cytokine-mediated killing (TRUCKs), which induce interleukin (IL)-12 release on target contact. TÜ165-based CAR-T cells (CAR-Ts) and TRUCKs with inducible IL-12 expression in an all-in-one construct were generated. Functionality of the engineered cells was assessed in co-cultures with EBNA-3C-peptide-loaded, HLA-B*35-expressing K562 cells and EBV-infected B cells as PTLD model. IL-12, secreted by TRUCKs on target contact, was further tested for its chemoattractive and activating potential towards monocytes and natural killer (NK) cells. After co-cultivation with EBV target cells, TÜ165 CAR-Ts and TRUCKs showed an increased activation marker expression (CD137, CD25) and release of proinflammatory cytokines (interferon-γ and tumor necrosis factor-α). Moreover, TÜ165 CAR-Ts and TRUCKs released apoptosis-inducing mediators (granzyme B and perforin) and were capable to specifically lyse EBV-positive target cells. Live cell imaging revealed a specific attraction of TÜ165 CAR-Ts around EBNA-3C-peptide-loaded target cells. Of note, TÜ165 TRUCKs with inducible IL-12 showed highly improved effector functions and additionally led to recruitment of monocyte and NK cell lines. Our results demonstrate that TÜ165 CAR-Ts recognize EBV peptide/HLA complexes in a TCR-like manner and thereby allow for recognizing an intracellular EBV target. TÜ165 TRUCKs equipped with inducible IL-12 expression responded even more effectively and released IL-12 recruited additional immune cells which are generally missing in proximity of lymphoproliferation in immunocompromised PTLD patients. This suggests a new and promising strategy to specifically target EBV-infected cells while sparing and mobilizing healthy immune cells and thereby enable control of EBV-associated lymphoproliferation.
Sections du résumé
BACKGROUND
Immunosuppressive therapy or T-cell depletion in transplant patients can cause uncontrolled growth of Epstein-Barr virus (EBV)-infected B cells resulting in post-transplant lymphoproliferative disease (PTLD). Current treatment options do not distinguish between healthy and malignant B cells and are thereby often limited by severe side effects in the already immunocompromised patients. To specifically target EBV-infected B cells, we developed a novel peptide-selective chimeric antigen receptor (CAR) based on the monoclonal antibody TÜ165 which recognizes an Epstein-Barr nuclear antigen (EBNA)-3C-derived peptide in HLA-B*35 context in a T-cell receptor (TCR)-like manner. In order to attract additional immune cells to proximity of PTLD cells, based on the TÜ165 CAR, we moreover generated T cells redirected for universal cytokine-mediated killing (TRUCKs), which induce interleukin (IL)-12 release on target contact.
METHODS
TÜ165-based CAR-T cells (CAR-Ts) and TRUCKs with inducible IL-12 expression in an all-in-one construct were generated. Functionality of the engineered cells was assessed in co-cultures with EBNA-3C-peptide-loaded, HLA-B*35-expressing K562 cells and EBV-infected B cells as PTLD model. IL-12, secreted by TRUCKs on target contact, was further tested for its chemoattractive and activating potential towards monocytes and natural killer (NK) cells.
RESULTS
After co-cultivation with EBV target cells, TÜ165 CAR-Ts and TRUCKs showed an increased activation marker expression (CD137, CD25) and release of proinflammatory cytokines (interferon-γ and tumor necrosis factor-α). Moreover, TÜ165 CAR-Ts and TRUCKs released apoptosis-inducing mediators (granzyme B and perforin) and were capable to specifically lyse EBV-positive target cells. Live cell imaging revealed a specific attraction of TÜ165 CAR-Ts around EBNA-3C-peptide-loaded target cells. Of note, TÜ165 TRUCKs with inducible IL-12 showed highly improved effector functions and additionally led to recruitment of monocyte and NK cell lines.
CONCLUSIONS
Our results demonstrate that TÜ165 CAR-Ts recognize EBV peptide/HLA complexes in a TCR-like manner and thereby allow for recognizing an intracellular EBV target. TÜ165 TRUCKs equipped with inducible IL-12 expression responded even more effectively and released IL-12 recruited additional immune cells which are generally missing in proximity of lymphoproliferation in immunocompromised PTLD patients. This suggests a new and promising strategy to specifically target EBV-infected cells while sparing and mobilizing healthy immune cells and thereby enable control of EBV-associated lymphoproliferation.
Identifiants
pubmed: 33127653
pii: jitc-2020-000736
doi: 10.1136/jitc-2020-000736
pmc: PMC7604878
pii:
doi:
Substances chimiques
EBNA-3C, epstein-barr virus
0
Epitopes
0
Epstein-Barr Virus Nuclear Antigens
0
HLA-B Antigens
0
Types de publication
Journal Article
Research Support, Non-U.S. Gov't
Langues
eng
Sous-ensembles de citation
IM
Informations de copyright
© Author(s) (or their employer(s)) 2020. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.
Déclaration de conflit d'intérêts
Competing interests: None declared.
Références
Sci Transl Med. 2013 Mar 13;5(176):176ra33
pubmed: 23486779
J Mol Med (Berl). 2014 Jul;92(7):735-41
pubmed: 24848996
Immunogenetics. 1981;13(4):359-65
pubmed: 6168583
Front Immunol. 2016 Sep 30;7:393
pubmed: 27746781
Cancer Cell Int. 2019 Jan 3;19:2
pubmed: 30622438
Gene Ther. 2000 Jun;7(12):1067-75
pubmed: 10871757
J Immunol. 2013 Nov 15;191(10):4989-95
pubmed: 24108698
Clin Cancer Res. 2013 Jun 15;19(12):3153-64
pubmed: 23620405
Proc Natl Acad Sci U S A. 2005 Jul 5;102(27):9571-6
pubmed: 15980149
Biol Blood Marrow Transplant. 2013 Oct;19(10):1480-92
pubmed: 23891747
Nat Rev Immunol. 2003 Feb;3(2):133-46
pubmed: 12563297
Cancers (Basel). 2020 Feb 06;12(2):
pubmed: 32041222
Front Immunol. 2018 Jun 27;9:1475
pubmed: 29997626
Expert Opin Biol Ther. 2015;15(8):1145-54
pubmed: 25985798
Blood. 2011 Apr 21;117(16):4262-72
pubmed: 21296998
Eur J Immunol. 1993 Mar;23(3):734-8
pubmed: 8449221
Oncoimmunology. 2017 Oct 11;7(1):e1378842
pubmed: 29296541
J Immunol. 2008 Oct 1;181(7):4874-82
pubmed: 18802091
J Immunol Methods. 2016 Mar;430:10-20
pubmed: 26780292
Nat Rev Clin Oncol. 2012 Sep;9(9):510-9
pubmed: 22801669
Mol Ther Oncolytics. 2017 Jan 11;3:1-9
pubmed: 29675462
Mol Ther. 2011 Apr;19(4):751-9
pubmed: 21285960
Leukemia. 2017 Aug;31(8):1788-1797
pubmed: 27924074
Pathogens. 2018 Apr 13;7(2):
pubmed: 29652813
Am J Transplant. 2011 Feb;11(2):336-47
pubmed: 21219573
Nucleic Acids Res. 2020 Jul 2;48(W1):W449-W454
pubmed: 32406916
J Biol Chem. 2005 Jan 28;280(4):2972-80
pubmed: 15537658
J Exp Med. 1986 Jul 1;164(1):196-210
pubmed: 3014034
J Clin Oncol. 2017 Feb 10;35(5):536-543
pubmed: 27992268
JCI Insight. 2018 Feb 22;3(4):
pubmed: 29467338
J Immunol. 2014 Dec 1;193(11):5733-43
pubmed: 25362181
Mol Ther. 2019 Feb 6;27(2):287-299
pubmed: 30573301
Sci Signal. 2018 Aug 21;11(544):
pubmed: 30131370
Sci Rep. 2014 Jan 06;4:3571
pubmed: 24389689
Clin Cancer Res. 2004 Apr 15;10(8):2626-35
pubmed: 15102664
Leukemia. 2015 Nov;29(11):2238-47
pubmed: 25987253
J Clin Invest. 2008 Jan;118(1):294-305
pubmed: 18060041
Nat Commun. 2020 Jan 10;11(1):219
pubmed: 31924795
Cancer Immunol Res. 2015 Feb;3(2):125-35
pubmed: 25212991
Proc Natl Acad Sci U S A. 1996 Mar 5;93(5):1820-4
pubmed: 8700842
Clin J Oncol Nurs. 2019 Apr 1;23(2):6-12
pubmed: 30880819
J Mol Biol. 1999 Jan 15;285(2):645-53
pubmed: 9878435
Proc Natl Acad Sci U S A. 2000 Jul 5;97(14):7969-74
pubmed: 10884427
Cancer Gene Ther. 2004 Feb;11(2):81-91
pubmed: 14685154
Cell Commun Signal. 2017 Jan 5;15(1):1
pubmed: 28073373
Front Immunol. 2017 Nov 24;8:1658
pubmed: 29225606
Blood. 2016 Sep 8;128(10):1396-407
pubmed: 27338099
Trends Immunol. 2015 Aug;36(8):494-502
pubmed: 26169254
J Hematol Oncol. 2018 Nov 27;11(1):132
pubmed: 30482221
Nucleic Acids Res. 2020 Jan 8;48(D1):D783-D788
pubmed: 31722398
J Transl Med. 2014 Dec 16;12:336
pubmed: 25510656
Cancer Lett. 2015 Dec 1;369(1):37-44
pubmed: 26279520
Proc Natl Acad Sci U S A. 2009 Apr 7;106(14):5784-8
pubmed: 19307587
Hum Gene Ther. 2017 Oct;28(10):914-925
pubmed: 28847167
Immunol Rev. 2014 Jan;257(1):83-90
pubmed: 24329791
Proc Natl Acad Sci U S A. 1993 Jan 15;90(2):720-4
pubmed: 8421711
Am J Transplant. 2006 Mar;6(3):569-76
pubmed: 16468968
Cancer Res. 2011 Sep 1;71(17):5697-706
pubmed: 21742772
Leuk Lymphoma. 2013 Nov;54(11):2433-40
pubmed: 23442063
Cancer Discov. 2013 Apr;3(4):388-98
pubmed: 23550147