Original Article

Integrated genomic analysis identifies deregulated JAK/STAT-MYC-biosynthesis axis in aggressive NK-cell leukemia

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Abstract

Aggressive NK-cell leukemia (ANKL) is a rare form of NK cell neoplasm that is more prevalent among people from Asia and Central and South America. Patients usually die within days to months, even after receiving prompt therapeutic management. Here we performed the first comprehensive study of ANKL by integrating whole genome, transcriptome and targeted sequencing, cytokine array as well as functional assays. Mutations in the JAK-STAT pathway were identified in 48% (14/29) of ANKL patients, while the extracellular STAT3 stimulator IL10 was elevated by an average of 56-fold (P < 0.0001) in the plasma of all patients examined. Additional frequently mutated genes included TP53 (34%), TET2 (28%), CREBBP (21%) and MLL2 (21%). Patient NK leukemia cells showed prominent activation of STAT3 phosphorylation, MYC expression and transcriptional activities in multiple metabolic pathways. Functionally, STAT3 activation and MYC expression were critical for the proliferation and survival of ANKL cells. STAT signaling regulated the MYC transcription program, and both STAT signaling and MYC transcription were required to maintain the activation of nucleotide synthesis and glycolysis. Collectively, the JAK-STAT pathway represents a major target for genomic alterations and IL10 stimulation in ANKL. This newly discovered JAK/STAT-MYC-biosynthesis axis may provide opportunities for the development of novel therapeutic strategies in treating this subtype of leukemia.

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Acknowledgements

We thank all the faculties and staffs in the Clinical and Laboratory Unit of the Department of Hematology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology for their clinical and technical support; the Core Genomic Facility of Beijing Institute of Genomics, Chinese Academy of Sciences for the construction and sequencing of libraries; Drs Qing Li and Jiguang Wang for critical reading and valuable comments on the manuscript; Dr Kai Fu for providing the cell line YT. This study was supported by the National Natural Science Foundation of China (81570196 to JZ, 81425003 to QW, 81670152 to Liang H, 81600120 to NW, 81300410 to DW, 81500100 to YL and 81400122 to KZ), the National Key Basic Research Program of China (2014CB542001 to QW), the Key Program of the National Natural Science Foundation of China (81230052 to JZ), the Key Research Program of the Chinese Academy of Sciences (Precious Medicine Research in Chinese Population; KJZD-EW-L14-3 to QW), and the National High Technology Research and Development Program of China (863 program; 2012AA02A507 to JZ and 2014AA020532 to Liang H).

Author information

Author notes

    • Liang Huang
    • , Dan Liu
    • , Na Wang
    • , Shaoping Ling
    •  & Yuting Tang

    These five authors contributed equally to this work.

Affiliations

  1. Department of Hematology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei 430030, China

    • Liang Huang
    • , Na Wang
    • , Yuting Tang
    • , Hui Luo
    • , Xuelian Hu
    • , Lingshuang Sheng
    • , Lijun Zhu
    • , Di Wang
    • , Yi Luo
    • , Zhen Shang
    • , Min Xiao
    • , Xia Mao
    • , Kuangguo Zhou
    •  & Jianfeng Zhou
  2. Key Laboratory of Genomic and Precision Medicine, Collaborative Innovation Center of Genetics and Development, Beijing Institute of Genomics, Chinese Academy of Sciences, Beijing 100101, China

    • Dan Liu
    • , Shaoping Ling
    • , Jun Wu
    • , Lingtong Hao
    • , Lihua Cao
    • , Lili Dong
    • , Xinchang Zheng
    • , Pinpin Sui
    • , Jianlin He
    • , Shanlan Mo
    • , Jin Yan
    • , Xin Liu
    •  & Qian-fei Wang
  3. Department of Pathology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei 430030, China

    • Qilin Ao
  4. Institute of Hematology and Blood Diseases Hospital, Chinese Academy of Medical Sciences & Peking Union Medical College, Tianjin 300020, China

    • Lugui Qiu
    • , Jianfeng Zhou
    •  & Qian-fei Wang
  5. University of Chinese Academy of Sciences, Beijing 100049, China

    • Lingtong Hao
    • , Xinchang Zheng
    • , Pinpin Sui
    • , Shanlan Mo
    • , Xin Liu
    •  & Qian-fei Wang
  6. Department of Hematology, Nanfang Hospital, Southern Medical University, Guangzhou, Guangdong 510515, China

    • Hongsheng Zhou
    •  & Qifa Liu
  7. Department of Hematology, Peking University Shenzhen Hospital, Shenzhen, Guangdong 518036, China

    • Hongyu Zhang
  8. Department of Hematology, the First Affiliated Hospital of Nanjing Medical University and Jiangsu Province Hospital, Nanjing, Jiangsu 210029, China

    • Jianyong Li
  9. Department of Hematology, the First Affiliated Hospital, Zhejiang University College of Medicine, Hangzhou, Zhejiang 310003, China

    • Jie Jin
  10. Department of Hematology, Beijing Friendship Hospital, Capital Medical University, Beijing 100050, China

    • Li Fu
  11. Shanghai Institute of Hematology, State Key Laboratory of Medical Genomics, Shanghai Rui Jin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China

    • Weili Zhao
  12. Department of Hematology, Southwest Hospital, Third Military Medical University, Chongqing 400038, China

    • Jieping Chen
  13. Department of Hematology, Guangdong General Hospital and Guangdong Academy of Medical Sciences, Guangzhou, Guangdong 510080, China

    • Xin Du
  14. Medical Research Institute, Wuhan University, Wuhan, Hubei 430071, China

    • Guoliang Qing
    •  & Hudan Liu
  15. Division of Experimental Hematology and Cancer Biology, Cincinnati Children's Hospital Medical Center, Cincinnati, OH 45229, USA

    • Gang Huang
  16. Division of Pathology, Cincinnati Children's Hospital Medical Center, Cincinnati, OH 45229, USA

    • Gang Huang
  17. Genome Wisdom Inc., Beijing 100195, China

    • Shaoping Ling
    • , Lingtong Hao
    •  & Lihua Cao
  18. Department of Obstetrics and Gynecology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei 430030, China

    • Ding Ma
  19. Cancer Biology Research Center, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei 430030, China

    • Ding Ma
    •  & Jianfeng Zhou

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Corresponding authors

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Supplementary information

PDF files

  1. 1.

    Supplementary information, Figure S1

    Strategies for the integrative analysis of ANKLs.

  2. 2.

    Supplementary information, Figure S2

    Isolation of granulocytes, leukemia NK cells and normal NK cells.

  3. 3.

    Supplementary information, Figure S3

    Frequency of somatic SNV of ANKL and 40 additional cancer types.

  4. 4.

    Supplementary information, Figure S4

    EBV reads number of leukemia NK cells and granulocytes from 8 ANKL patients.

  5. 5.

    Supplementary information, Figure S5

    KEGG pathway enrichment analysis of protein-altering somatic SNVs.

  6. 6.

    Supplementary information, Figure S6

    Functional enrichment analysis of genomic alterations with HotNet2.

  7. 7.

    Supplementary information, Figure S7

    STAT3 and STAT5B mutations identified in ANKL patients.

  8. 8.

    Supplementary information, Figure S8

    TET2 mutations identified in ANKL patients.

  9. 9.

    Supplementary information, Figure S9

    Semi-quantitative immunoreactivity histological scores of MYC staining in bone marrow biopsy of ANKL and control samples.

  10. 10.

    Supplementary information, Figure S10

    Functional enrichment map for MYC-signature genes that were upregulated in ANKLs compared to healthy donors.

  11. 11.

    Supplementary information, Figure S11

    The effect of IL10 or a STAT3 inhibitor (Stattic) on the apoptosis of ANKL cell lines.

  12. 12.

    Supplementary information, Figure S12

    The effect of IL10 and the STAT3 inhibitor (Stattic) on the mRNA expression of MYC in ANKL cell lines.

  13. 13.

    Supplementary information, Figure S13

    The rate of EdU incorporation in JQ1-treated ANKL cell lines.

  14. 14.

    Supplementary information, Figure S14

    Gene-set enrichment analysis (GSEA) of known MYC and STAT3 signatures.

  15. 15.

    Supplementary information, Figure S15

    Enrichment of metabolic pathways in primary ANKL leukemia cells and ANKL cell lines.

  16. 16.

    Supplementary information, Figure S16

    The effect of STAT3 Y640F mutant on the phosphorylation of STAT3 and mRNA expression of MYC target gene.

  17. 17.

    Supplementary information, Figure S17

    The effect of IL10 on the mRNA expression of MYC and MYC target genes in KHYG-1 cell line transfected with STAT3 Y640F mutant.

  18. 18.

    Supplementary information, Figure S18

    The effect of IL10 on STAT3 phosphorylation, MYC expressionand the proliferation of STAT3 Y640F-mutant ANKL cell line YT.

  19. 19.

    Supplementary information, Figure S19

    Quantitative analysis of the Epstein-Barr virus (EBV) load in ANKL patients and healthy donors with whole-transcriptome sequencing (WTS) data.

  20. 20.

    Supplementary information, Figure S20

    Expression of EBV-encoded small RNAs in primary ANKL leukemia cells.

  21. 21.

    Supplementary information, Figure S21

    A large gain across MYC and an inter-chromosomal translocation detected by GVC-CNV and GVC-SV in ANKL No.19.

  22. 22.

    Supplementary information, Table S1

    Patient characteristics of ANKL.

  23. 23.

    Supplementary information, Table S2

    SNVs identified in ANKL NK leukemia cells.

  24. 24.

    Supplementary information, Table S3

    CNVs identified in ANKL NK leukemia cells.

  25. 25.

    Supplementary information, Table S4

    SVs identified in ANKL NK leukemia cells.

  26. 26.

    Supplementary information, Table S5

    AmpliSeq targeted sequencing results of ANKLs.

  27. 27.

    Supplementary information, Table S6

    Comparison of the mutational profiles between ANKL and NKTCL10,11.

  28. 28.

    Supplementary information, Table S7

    H-score of phosphorylated STAT3 and corresponding STAT mutation status of ANKL cases.

  29. 29.

    Supplementary information, Table S8

    Differentially upregulated genes in ANKL leukemia cells in comparison with normal controls.

  30. 30.

    Supplementary information, Table S9

    Differentially downregulated genes in ANKL leukemia cells in comparison with normal controls.

  31. 31.

    Supplementary information, Table S10

    KEGG pathway enrichment analysis of upregulated genes in ANKL leukemia cells.

  32. 32.

    Supplementary information, Table S11

    KEGG pathway enrichment analysis of downregulated genes in ANKL leukemia cells.

  33. 33.

    Supplementary information, Table S12

    IPA metabolic pathways analysis of ANKL and tumors that have known metabolic features.

  34. 34.

    Supplementary information, Table S13

    Sequencing depth information of 8 WGS ANKL patients.

  35. 35.

    Supplementary information, Table S14

    Primers for quantitative RT-PCR

  36. 36.

    Supplementary information, Data S1

    Materials and methods

(Supplementary information is linked to the online version of the paper on the Cell Research website.)