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Non-Hodgkin Lymphoma
Genetic manipulation of primary human natural killer cells to investigate the functional and oncogenic roles of PRDM1
Gehong Dong,1,2* Yuping Li,1* Logan Lee,1 Xuxiang Liu,1 Yunfei Shi,1,3 Xiaoqian Liu,1,4 Alyssa Bouska,5 Qiang Gong,1 Lingbo Kong,1 Jinhui Wang,6 Chih-Hong Lou,7 Timothy W. McKeithan,1 Javeed Iqbal5 and Wing C. Chan1
1Department of Pathology, City of Hope National Medical Center, Duarte, CA, USA; 2Department of Pathology, Beijing Tiantan Hospital, Capital Medical University, Beijing, China; 3Department of Pathology, Peking University Cancer Hospital & Institute, Key Laboratory of Carcinogenesis and Translational Research (Ministry of Education), Beijing, China; 4Department of Hematology, Affiliated Yantai Yuhuangding Hospital, Qingdao University, Yantai, Shandong, China; 5Pathology and Microbiology, University of Nebraska Medical Center, Omaha, NE, USA; 6Department of Molecular and Cellular Biology, City of Hope, Duarte, CA, USA and 7The Gene Editing and Viral Vector Core, Department of Shared Resources, Beckman Research Institute of City of Hope, Duarte, CA, USA
*GD and YL contributed equally as co-first authors.
ABSTRACT
Extra-nodal natural killer (NK)/T-cell lymphoma, nasal type (ENKTCL) is a highly aggressive lymphoma, in which the tumor suppressor gene PRDM1 is frequently lost or inactivated. We employed two different CRISPR/Cas9 approaches to generate PRDM1-/- primary NK cells to study the role of this gene in NK-cell homeostasis. PRDM1-/- NK cells showed a marked increase in cloning efficiency, high- er proliferation rate and less apoptosis compared with their wild-type counterparts. Gene expression profiling demonstrated a marked enrich- ment in pathways associated with proliferation, cell cycle, MYC, MYB and TCR/NK signaling in PRDM1-/- NK cells, but pathways associated with normal cellular functions including cytotoxic functions were down- regulated, suggesting that the loss of PRDM1 shifted NK cells toward proliferation and survival rather than the performance of their normal functions. We were also able to further modify a PRDM1-deleted clone to introduce heterozygous deletions of common tumor suppressor genes in ENKTCL such as TP53, DDX3X, and PTPN6. We established an in vitro model to elucidate the major pathways through which PRDM1 mediates its homeostatic control of NK cells. This approach can be applied to the study of other relevant genetic lesions and oncogenic collaborations in lymphoma pathogenesis.
Introduction
Extra-nodal natural killer (NK)/T-cell lymphoma, nasal type (ENKTCL) is a highly aggressive lymphoma that is consistently associated with Epstein-Barr virus (EBV) infection and predominantly affects middle-aged men in Asia and Central and South America.1,2 It typically presents as tumors or destructive lesions in the nasal cavity, maxillary sinuses or palate. Despite a localized presentation in most patients, it tends to relapse locally or at other extra-nodal sites, such as the skin, and the 5-year overall survival of affected individuals is 40-50% with current therapeutic regimens.2,3 About 80-90% of ENKTCL originate from the NK-cell lineage with the rest of cases derived from T cells. Regardless of the cell of origin, the pathology, clinical behavior and treat- ment are similar. Aggressive NK-cell leukemia, also EBV-associated and derived from NK cells, is regarded as the leukemic form of ENKTCL.4
Our previous genomic analysis of ENKTCL,5,6 including identification of copy number abnormalities, mutation analysis, and DNA methylation studies, suggested that PRDM1, located in 6q21, is a tumor suppressor gene that is frequently inactivat-
Ferrata Storti Foundation
Haematologica 2021 Volume 106(9):2427-2438
Correspondence:
WING C. (JOHN) CHAN
jochan@coh.org
Received: April 8, 2020. Accepted: July 30, 2020. Pre-published: July 30, 2020.
https://doi.org/10.3324/haematol.2020.254276 ©2021 Ferrata Storti Foundation
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