Letter | Published:

Cyclin D–CDK4 kinase destabilizes PD-L1 via cullin 3–SPOP to control cancer immune surveillance

Nature volume 553, pages 9195 (04 January 2018) | Download Citation


Treatments that target immune checkpoints, such as the one mediated by programmed cell death protein 1 (PD-1) and its ligand PD-L1, have been approved for treating human cancers with durable clinical benefit1,2. However, many patients with cancer fail to respond to compounds that target the PD-1 and PD-L1 interaction, and the underlying mechanism(s) is not well understood3,4,5. Recent studies revealed that response to PD-1–PD-L1 blockade might correlate with PD-L1 expression levels in tumour cells6,7. Hence, it is important to understand the mechanistic pathways that control PD-L1 protein expression and stability, which can offer a molecular basis to improve the clinical response rate and efficacy of PD-1–PD-L1 blockade in patients with cancer. Here we show that PD-L1 protein abundance is regulated by cyclin D–CDK4 and the cullin 3–SPOP E3 ligase via proteasome-mediated degradation. Inhibition of CDK4 and CDK6 (hereafter CDK4/6) in vivo increases PD-L1 protein levels by impeding cyclin D–CDK4-mediated phosphorylation of speckle-type POZ protein (SPOP) and thereby promoting SPOP degradation by the anaphase-promoting complex activator FZR1. Loss-of-function mutations in SPOP compromise ubiquitination-mediated PD-L1 degradation, leading to increased PD-L1 levels and reduced numbers of tumour-infiltrating lymphocytes in mouse tumours and in primary human prostate cancer specimens. Notably, combining CDK4/6 inhibitor treatment with anti-PD-1 immunotherapy enhances tumour regression and markedly improves overall survival rates in mouse tumour models. Our study uncovers a novel molecular mechanism for regulating PD-L1 protein stability by a cell cycle kinase and reveals the potential for using combination treatment with CDK4/6 inhibitors and PD-1–PD-L1 immune checkpoint blockade to enhance therapeutic efficacy for human cancers.

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  1. 1.

    , & PD-L1 (B7-H1) and PD-1 pathway blockade for cancer therapy: Mechanisms, response biomarkers, and combinations. Sci. Transl. Med. 8, 328rv4 (2016)

  2. 2.

    Molecular and biochemical aspects of the PD-1 checkpoint pathway. N. Engl. J. Med. 375, 1767–1778 (2016)

  3. 3.

    et al. Prospects for combining targeted and conventional cancer therapy with immunotherapy. Nat. Rev. Cancer 17, 286–301 (2017)

  4. 4.

    & The future of immune checkpoint therapy. Science 348, 56–61 (2015)

  5. 5.

    , & Combination cancer immunotherapy and new immunomodulatory targets. Nat. Rev. Drug Discov. 14, 561–584 (2015)

  6. 6.

    et al. Predictive correlates of response to the anti-PD-L1 antibody MPDL3280A in cancer patients. Nature 515, 563–567 (2014)

  7. 7.

    et al. Involvement of PD-L1 on tumor cells in the escape from host immune system and tumor immunotherapy by PD-L1 blockade. Proc. Natl Acad. Sci. USA 99, 12293–12297 (2002)

  8. 8.

    & Cell cycle proteins as promising targets in cancer therapy. Nat. Rev. Cancer 17, 93–115 (2017)

  9. 9.

    et al. MYC regulates the antitumor immune response through CD47 and PD-L1. Science 352, 227–231 (2016)

  10. 10.

    et al. Cdk5 disruption attenuates tumor PD-L1 expression and promotes antitumor immunity. Science 353, 399–403 (2016)

  11. 11.

    et al. Glycosylation and stabilization of programmed death ligand-1 suppresses T-cell activity. Nat. Commun. 7, 12632 (2016)

  12. 12.

    et al. Deubiquitination and stabilization of PD-L1 by CSN5. Cancer Cell 30, 925–939 (2016)

  13. 13.

    & Taxol stabilizes microtubules in mouse fibroblast cells. Proc. Natl Acad. Sci. USA 77, 1561–1565 (1980)

  14. 14.

    & Mammalian cyclin-dependent kinases. Trends Biochem. Sci. 30, 630–641 (2005)

  15. 15.

    , & Non-canonical functions of cell cycle cyclins and cyclin-dependent kinases. Nat. Rev. Mol. Cell Biol. 17, 280–292 (2016)

  16. 16.

    et al. CDK6 (PLSTIRE) and CDK4 (PSK-J3) are a distinct subset of the cyclin-dependent kinases that associate with cyclin D1. Oncogene 9, 71–79 (1994)

  17. 17.

    ., ., ., & Cyclin E/cdk2 and cyclin A/cdk2 kinases associate with p107 and E2F in a temporally distinct manner. Genes Dev. 6, 1874–1885 (1992)

  18. 18.

    et al. Preferences for phosphorylation sites in the retinoblastoma protein of D-type cyclin-dependent kinases, Cdk4 and Cdk6, in vitro. J. Biochem. 137, 381–386 (2005)

  19. 19.

    et al. Specific inhibition of cyclin-dependent kinase 4/6 by PD 0332991 and associated antitumor activity in human tumor xenografts. Mol. Cancer Ther. 3, 1427–1438 (2004)

  20. 20.

    , , & Lack of cyclin D–Cdk complexes in Rb-negative cells correlates with high levels of p16INK4/MTS1 tumour suppressor gene product. EMBO J. 14, 503–511 (1995)

  21. 21.

    et al. Retinoblastoma-protein-dependent cell-cycle inhibition by the tumour suppressor p16. Nature 375, 503–506 (1995)

  22. 22.

    et al. An inhibitor of NEDD8-activating enzyme as a new approach to treat cancer. Nature 458, 732–736 (2009)

  23. 23.

    , & The emerging family of CULLIN3-RING ubiquitin ligases (CRL3s): cellular functions and disease implications. EMBO J. 32, 2307–2320 (2013)

  24. 24.

    et al. Exome sequencing identifies recurrent SPOP, FOXA1 and MED12 mutations in prostate cancer. Nat. Genet. 44, 685–689 (2012)

  25. 25.

    Cancer Genome Atlas Research. The molecular taxonomy of primary prostate cancer. Cell 163, 1011–1025 (2015)

  26. 26.

    et al. Integrated molecular analysis of clear-cell renal cell carcinoma. Nat. Genet. 45, 860–867 (2013)

  27. 27.

    et al. Genome-wide association study in Chinese men identifies two new prostate cancer risk loci at 9q31.2 and 19q13.4. Nat. Genet. 44, 1231–1235 (2012)

  28. 28.

    et al. SPOP promotes ubiquitination and degradation of the ERG oncoprotein to suppress prostate cancer progression. Mol. Cell 59, 917–930 (2015)

  29. 29.

    et al. Structures of APC/CCdh1 with substrates identify Cdh1 and Apc10 as the D-box co-receptor. Nature 470, 274–278 (2011)

  30. 30.

    et al. CDK4/6 inhibition triggers anti-tumour immunity. Nature 548, 471–475 (2017)

  31. 31.

    et al. Prostate cancer-associated SPOP mutations confer resistance to BET inhibitors through stabilization of BRD4. Nat. Med. 23, 1063–1071 (2017)

  32. 32.

    et al. The APC/C E3 ligase complex activator FZR1 restricts BRAF oncogenic function. Cancer Discov. 7, 424–441 (2017)

  33. 33.

    et al. Phosphorylation by Akt1 promotes cytoplasmic localization of Skp2 and impairs APCCdh1-mediated Skp2 destruction. Nat. Cell Biol. 11, 397–408 (2009)

  34. 34.

    et al. Requirement for CDK4 kinase function in breast cancer. Cancer Cell 9, 23–32 (2006)

  35. 35.

    et al. Requirement for cyclin D3 in lymphocyte development and T cell leukemias. Cancer Cell 4, 451–461 (2003)

  36. 36.

    et al. Programmed cell death 1 ligand 1 and tumor-infiltrating CD8+ T lymphocytes are prognostic factors of human ovarian cancer. Proc. Natl Acad. Sci. USA 104, 3360–3365 (2007)

  37. 37.

    et al. The metabolic function of cyclin D3-CDK6 kinase in cancer cell survival. Nature 546, 426–430 (2017)

  38. 38.

    et al. APCCdc20 suppresses apoptosis through targeting Bim for ubiquitination and destruction. Dev. Cell 29, 377–391 (2014)

  39. 39.

    et al. Mouse development and cell proliferation in the absence of D-cyclins. Cell 118, 477–491 (2004)

  40. 40.

    et al. The requirement for cyclin D function in tumor maintenance. Cancer Cell 22, 438–451 (2012)

  41. 41.

    et al. Cyclin D1 provides a link between development and oncogenesis in the retina and breast. Cell 82, 621–630 (1995)

  42. 42.

    et al. Specific protection against breast cancers by cyclin D1 ablation. Nature 411, 1017–1021 (2001)

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We thank members of the Wei, Freeman, Sicinski, and Pandolfi laboratories for discussions. J.Z. is supported by the career transition award (1K99CA212292-01). W.W. is a Leukemia & Lymphoma Society research scholar. This work was supported in part by the National Institutes of Health (NIH) grants GM094777 and CA177910 (to W.W.), P01 CA080111, R01 CA202634 and R01 CA132740 (to P.S.), and P50CA101942 (to G.J.F).

Author information

Author notes

    • Jinfang Zhang
    • , Xia Bu
    • , Haizhen Wang
    • , Gordon J. Freeman
    • , Piotr Sicinski
    •  & Wenyi Wei

    These authors contributed equally to this work.

    • Gordon J. Freeman
    • , Piotr Sicinski
    •  & Wenyi Wei

    These authors jointly supervised this work.


  1. Department of Pathology, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, Massachusetts 02215, USA

    • Jinfang Zhang
    • , Naoe Taira Nihira
    • , Yuyong Tan
    • , Yanpeng Ci
    • , Fei Wu
    • , Xiangpeng Dai
    • , Jianping Guo
    • , Yu-Han Huang
    •  & Wenyi Wei
  2. Department of Medical Oncology, Dana-Farber Cancer Institute, Harvard Medical School, Boston, Massachusetts 02215, USA

    • Xia Bu
    •  & Gordon J. Freeman
  3. Department of Cancer Biology, Dana-Farber Cancer Institute and Department of Genetics, Harvard Medical School, Boston, Massachusetts 02215, USA

    • Haizhen Wang
    • , Yan Geng
    • , Caoqi Fan
    •  & Piotr Sicinski
  4. Department of Urology, Shanghai Changhai Hospital, Second Military Medical University, Shanghai 200433, China

    • Yasheng Zhu
    • , Shancheng Ren
    •  & Yinghao Sun
  5. Department of Gastroenterology, the Second Xiangya Hospital of Central South University, Changsha 410011, China

    • Yuyong Tan
  6. School of Life Science and Technology, Harbin Institute of Technology, Harbin 150001, China

    • Yanpeng Ci
  7. Department of Urology, Huashan Hospital, Fudan University, Shanghai 200040, China.

    • Fei Wu
  8. Peking-Tsinghua Center for Life Sciences, Academy for Advanced Interdisciplinary Studies, School of Life Sciences, Peking University, Beijing 100871, China

    • Caoqi Fan


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J.Z., X.B. and H.W. performed most of the experiments with assistance from Y.Z., Y.G., N.T.N., Y.T., Y.C., F.W., X.D., J.G., Y.H., C.F., S.R. and Y.S. Y.Z., S.R., and Y.S. performed immunohistochemistry for human prostate cancer samples. Y.G., Y.T. and Y.C. helped with mice work. J.Z., X.B., H.W., G.J.F., P.S. and W.W. designed the experiments. G.J.F., P.S. and W.W. supervised the study. J.Z. and W.W. wrote the manuscript with help from X.B., H.W., P.S. and G.J.F. All authors commented on the manuscript.

Competing interests

G.J.F. has patents and pending royalties on the PD-1 pathway from Roche, Merck, Bristol-Myers-Squibb, EMD-Serono, Boehringer-Ingelheim, AstraZeneca, DAKO and Novartis. G.J.F. has served on advisory boards for CoStim, Novartis, Roche, Eli Lilly, Bristol-Myers-Squibb, Seattle Genetics, Bethyl Laboratories, Xios, and Quiet. P.S. is a consultant and a recipient of a research grant from Novartis. No potential conflicts of interests were disclosed by other authors.

Corresponding authors

Correspondence to Gordon J. Freeman or Piotr Sicinski or Wenyi Wei.

Reviewer Information Nature thanks J. Bartek, C. Klebanoff and S. Ogawa for their contribution to the peer review of this work.

Publisher's note: Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

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    Supplementary Figure 1

    This file contains the source data for gels in Figures 1-4 and Extended Data Figures 1-9.

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