Article

DKK2 imparts tumor immunity evasion through β-catenin-independent suppression of cytotoxic immune-cell activation

Received:
Accepted:
Published online:

Abstract

Immunotherapy offers new options for cancer treatment, but efficacy varies across cancer types. Colorectal cancers (CRCs) are largely refractory to immune-checkpoint blockade, which suggests the presence of yet uncharacterized immune-suppressive mechanisms. Here we report that the loss of adenomatosis polyposis coli (APC) in intestinal tumor cells or of the tumor suppressor PTEN in melanoma cells upregulates the expression of Dickkopf-related protein 2 (DKK2), which, together with its receptor LRP5, provides an unconventional mechanism for tumor immune evasion. DKK2 secreted by tumor cells acts on cytotoxic lymphocytes, inhibiting STAT5 signaling by impeding STAT5 nuclear localization via LRP5, but independently of LRP6 and the Wnt–β-catenin pathway. Genetic or antibody-mediated ablation of DKK2 activates natural killer (NK) cells and CD8+ T cells in tumors, impedes tumor progression, and enhances the effects of PD-1 blockade. Thus, we have identified a previously unknown tumor immune-suppressive mechanism and immunotherapeutic targets particularly relevant for CRCs and a subset of melanomas.

  • Subscribe to Nature Medicine for full access:

    $225

    Subscribe

Additional access options:

Already a subscriber?  Log in  now or  Register  for online access.

Accessions

Primary accessions

Gene Expression Omnibus

References

  1. 1.

    & Oncology meets immunology: the cancer-immunity cycle. Immunity 39, 1–10 (2013).

  2. 2.

    The blockade of immune checkpoints in cancer immunotherapy. Nat. Rev. Cancer 12, 252–264 (2012).

  3. 3.

    , & Immune checkpoint blockade in cancer therapy. J. Clin. Oncol. 33, 1974–1982 (2015).

  4. 4.

    , & Neutralizing tumor-promoting chronic inflammation: a magic bullet? Science 339, 286–291 (2013).

  5. 5.

    , & Immune checkpoint blockade: a common denominator approach to cancer therapy. Cancer Cell 27, 450–461 (2015).

  6. 6.

    & The basis of oncoimmunology. Cell 164, 1233–1247 (2016).

  7. 7.

    et al. Safety, activity, and immune correlates of anti-PD-1 antibody in cancer. N. Engl. J. Med. 366, 2443–2454 (2012).

  8. 8.

    et al. Safety and activity of anti-PD-L1 antibody in patients with advanced cancer. N. Engl. J. Med. 366, 2455–2465 (2012).

  9. 9.

    et al. Phase II study of the anti-cytotoxic T-lymphocyte-associated antigen 4 monoclonal antibody, tremelimumab, in patients with refractory metastatic colorectal cancer. J. Clin. Oncol. 28, 3485–3490 (2010).

  10. 10.

    , , & Primary, adaptive, and acquired resistance to cancer immunotherapy. Cell 168, 707–723 (2017).

  11. 11.

    The biology of interleukin-2 and interleukin-15: implications for cancer therapy and vaccine design. Nat. Rev. Immunol. 6, 595–601 (2006).

  12. 12.

    , & Molecular pathways: interleukin-15 signaling in health and in cancer. Clin. Cancer Res. 20, 2044–2050 (2014).

  13. 13.

    et al. The transcription factor T-bet is induced by IL-15 and thymic agonist selection and controls CD8αα(+) intraepithelial lymphocyte development. Immunity 41, 230–243 (2014).

  14. 14.

    , & The light and dark sides of intestinal intraepithelial lymphocytes. Nat. Rev. Immunol. 11, 445–456 (2011).

  15. 15.

    , & Interleukin 15 as a promising candidate for tumor immunotherapy. Cytokine Growth Factor Rev. 22, 99–108 (2011).

  16. 16.

    et al. Interleukin-7 and interleukin-15 for cancer. J. Cancer 5, 765–773 (2014).

  17. 17.

    et al. The cytotoxic T cell proteome and its shaping by the kinase mTOR. Nat. Immunol. 17, 104–112 (2016).

  18. 18.

    , , & The critical role of IL-15-PI3K-mTOR pathway in natural killer cell effector functions. Front. Immunol. 5, 187 (2014).

  19. 19.

    & The Wnt signaling pathway in development and disease. Annu. Rev. Cell Dev. Biol. 20, 781–810 (2004).

  20. 20.

    , , & WNT and beta-catenin signalling: diseases and therapies. Nat. Rev. Genet. 5, 691–701 (2004).

  21. 21.

    , & Wnt/beta-catenin signaling: components, mechanisms, and diseases. Dev. Cell 17, 9–26 (2009).

  22. 22.

    & Wnt/β-catenin signaling and disease. Cell 149, 1192–1205 (2012).

  23. 23.

    The complex world of WNT receptor signalling. Nat. Rev. Mol. Cell Biol. 13, 767–779 (2012).

  24. 24.

    Wnt signaling in cancer. Cold Spring Harb. Perspect. Biol. 4, a008052 (2012).

  25. 25.

    Function and biological roles of the Dickkopf family of Wnt modulators. Oncogene 25, 7469–7481 (2006).

  26. 26.

    , & The structural basis of DKK-mediated inhibition of Wnt/LRP signaling. Sci. Signal. 5, pe22 (2012).

  27. 27.

    , , & The canonical Wnt signaling antagonist DKK2 is an essential effector of PITX2 function during normal eye development. Dev. Biol. 317, 310–324 (2008).

  28. 28.

    et al. Dkk2 has a role in terminal osteoblast differentiation and mineralized matrix formation. Nat. Genet. 37, 945–952 (2005).

  29. 29.

    et al. Dkk2 plays an essential role in the corneal fate of the ocular surface epithelium. Development 133, 2149–2154 (2006).

  30. 30.

    et al. Chemical and genetic evidence for the involvement of Wnt antagonist Dickkopf2 in regulation of glucose metabolism. Proc. Natl. Acad. Sci. USA 109, 11402–11407 (2012).

  31. 31.

    , , & Mutual antagonism between dickkopf1 and dickkopf2 regulates Wnt/beta-catenin signalling. Curr. Biol. 10, 1611–1614 (2000).

  32. 32.

    , , , & Second cysteine-rich domain of Dickkopf-2 activates canonical Wnt signaling pathway via LRP-6 independently of dishevelled. J. Biol. Chem. 277, 5977–5981 (2002).

  33. 33.

    et al. Dickkopf-1 regulates gastrulation movements by coordinated modulation of Wnt/beta catenin and Wnt/PCP activities, through interaction with the Dally-like homolog Knypek. Genes Dev. 21, 465–480 (2007).

  34. 34.

    et al. Mutated KRAS results in overexpression of DUSP4, a MAP-kinase phosphatase, and SMYD3, a histone methyltransferase, in rectal carcinomas. Genes Chromosom. Cancer 49, 1024–1034 (2010).

  35. 35.

    et al. DICKKOPF-4 and -2 genes are upregulated in human colorectal cancer. Cancer Sci. 100, 1923–1930 (2009).

  36. 36.

    Cancer Genome Atlas Network. Comprehensive molecular characterization of human colon and rectal cancer. Nature 487, 330–337 (2012).

  37. 37.

    et al. Multiple intestinal neoplasia caused by a mutation in the murine homolog of the APC gene. Science 256, 668–670 (1992).

  38. 38.

    , & Cytotoxic and non-cytotoxic roles of the CTL/NK protease granzyme B. Immunol. Rev. 235, 105–116 (2010).

  39. 39.

    et al. Dickkopf-related protein 1 (Dkk1) regulates the accumulation and function of myeloid derived suppressor cells in cancer. J. Exp. Med. 213, 827–840 (2016).

  40. 40.

    et al. Metastatic latency and immune evasion through autocrine inhibition of WNT. Cell 165, 45–60 (2016).

  41. 41.

    et al. Jak1 has a dominant role over Jak3 in signal transduction through γc-containing cytokine receptors. Chem. Biol. 18, 314–323 (2011).

  42. 42.

    et al. Genomic and transcriptomic features of response to anti-PD-1 therapy in metastatic melanoma. Cell 165, 35–44 (2016).

  43. 43.

    et al. Loss of PTEN promotes resistance to T cell-mediated immunotherapy. Cancer Discov. 6, 202–216 (2016).

  44. 44.

    et al. sFRP2 in the aged microenvironment drives melanoma metastasis and therapy resistance. Nature 532, 250–254 (2016).

  45. 45.

    , & DKK1 antagonizes Wnt signaling without promotion of LRP6 internalization and degradation. J. Biol. Chem. 283, 21427–21432 (2008).

  46. 46.

    et al. Clathrin and AP2 are required for PtdIns(4,5)P2-mediated formation of LRP6 signalosomes. J. Cell Biol. 200, 419–428 (2013).

  47. 47.

    et al. STAT5 is a key regulator in NK cells and acts as a molecular switch from tumor surveillance to tumor promotion. Cancer Discov. 6, 414–429 (2016).

  48. 48.

    et al. Stat5b is essential for natural killer cell-mediated proliferation and cytolytic activity. J. Exp. Med. 188, 2067–2074 (1998).

  49. 49.

    et al. Stat5a and Stat5b proteins have essential and nonessential, or redundant, roles in cytokine responses. Cell 93, 841–850 (1998).

  50. 50.

    et al. A novel Ncr1-Cre mouse reveals the essential role of STAT5 for NK-cell survival and development. Blood 117, 1565–1573 (2011).

  51. 51.

    , & Wnt signaling induces matrix metalloproteinase expression and regulates T cell transmigration. Immunity 26, 227–239 (2007).

  52. 52.

    et al. Recognition of tumors by the innate immune system and natural killer cells. Adv. Immunol. 122, 91–128 (2014).

  53. 53.

    , , & NK cells regulating T cell responses: mechanisms and outcome. Trends Immunol. 36, 49–58 (2015).

  54. 54.

    et al. Lrp5 functions in bone to regulate bone mass. Nat. Med. 17, 684–691 (2011).

  55. 55.

    et al. Blockade of B7-H1 and PD-1 by monoclonal antibodies potentiates cancer therapeutic immunity. Cancer Res. 65, 1089–1096 (2005).

  56. 56.

    , & Cell surface markers in colorectal cancer prognosis. Int. J. Mol. Sci. 12, 78–113 (2010).

  57. 57.

    et al. Loss of Apc in vivo immediately perturbs Wnt signaling, differentiation, and migration. Genes Dev. 18, 1385–1390 (2004).

  58. 58.

    et al. Differentiation and reversal of malignant changes in colon cancer through PPARγ. Nat. Med. 4, 1046–1052 (1998).

  59. 59.

    & Stem cell dynamics in homeostasis and cancer of the intestine. Nat. Rev. Cancer 14, 468–480 (2014).

  60. 60.

    et al. Characterization of the Kremen-binding site on Dkk1 and elucidation of the role of Kremen in Dkk-mediated Wnt antagonism. J. Biol. Chem. 283, 23371–23375 (2008).

  61. 61.

    , , & The characterization of intraepithelial lymphocytes, lamina propria leukocytes, and isolated lymphoid follicles in the large intestine of mice infected with the intestinal nematode parasite Trichuris muris. J. Immunol. 175, 6713–6722 (2005).

  62. 62.

    et al. Small intestinal intraepithelial lymphocytes expressing CD8 and T cell receptor γδ are involved in bacterial clearance during Salmonella enterica serovar Typhimurium infection. Infect. Immun. 80, 565–574 (2012).

  63. 63.

    et al. Genome engineering using the CRISPR-Cas9 system. Nat. Protoc. 8, 2281–2308 (2013).

  64. 64.

    et al. Gene set enrichment analysis: a knowledge-based approach for interpreting genome-wide expression profiles. Proc. Natl. Acad. Sci. USA 102, 15545–15550 (2005).

Download references

Acknowledgements

We thank M. Orsulak for technical assistance and B. Williams (Van Andel Institute, Grand Rapids, Michigan, USA) for providing the LRP5/6 floxed mice. This work was supported by the NIH (grants GM112182 and CA214703 to D.W.), the Connecticut Bioscience Innovation Fund (to D.W.), NSFC (grant 31530094 to L.L.), the strategic priority research program of CAS (grant XDB19000000 to L.L.), and the CAS/SAFEA International Partnership Program for Creative Research Teams (to L.L. and D.W.).

Author information

Author notes

    • Qian Xiao
    • , Jibo Wu
    •  & Wei-Jia Wang

    These authors contributed equally to this work.

Affiliations

  1. Vascular Biology and Therapeutic Program and Department of Pharmacology, Yale School of Medicine, New Haven, Connecticut, USA.

    • Qian Xiao
    • , Wei-Jia Wang
    • , Yingxia Zheng
    • , Mahnaz Sahraei
    • , Wenwen Tang
    •  & Dianqing Wu
  2. State Key Laboratory of Molecular Biology, CAS Center for Excellence in Molecular Cell Science, Innovation Center for Cell Signaling Network, Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, University of Chinese Academy of Sciences, Shanghai, China.

    • Jibo Wu
    •  & Lin Li
  3. State Key Laboratory of Cell Biology, CAS Center for Excellence in Molecular Cell Science, Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences, University of Chinese Academy of Sciences, Shanghai, China.

    • Shiyang Chen
    •  & Jianfeng Chen
  4. Biostatistics Department, Yale University, New Haven, Connecticut, USA.

    • Xiaoqing Yu
  5. Departments of Dermatology and Pathology, Yale School of Medicine, New Haven, Connecticut, USA.

    • Katrina Meeth
    •  & Marcus Bosenberg
  6. Department of Immunobiology, Yale School of Medicine, New Haven, Connecticut, USA.

    • Alfred L M Bothwell
    • , Lieping Chen
    •  & Dianqing Wu
  7. Yale Cancer Center, Yale School of Medicine, New Haven, Connecticut, USA.

    • Alfred L M Bothwell
    • , Lieping Chen
    •  & Marcus Bosenberg
  8. Institute of Pharmacology and Toxicology, Department for Biomedical Sciences, University of Veterinary Medicine Vienna, Vienna, Austria.

    • Veronika Sexl
  9. AbMax, Beijing, China.

    • Le Sun

Authors

  1. Search for Qian Xiao in:

  2. Search for Jibo Wu in:

  3. Search for Wei-Jia Wang in:

  4. Search for Shiyang Chen in:

  5. Search for Yingxia Zheng in:

  6. Search for Xiaoqing Yu in:

  7. Search for Katrina Meeth in:

  8. Search for Mahnaz Sahraei in:

  9. Search for Alfred L M Bothwell in:

  10. Search for Lieping Chen in:

  11. Search for Marcus Bosenberg in:

  12. Search for Jianfeng Chen in:

  13. Search for Veronika Sexl in:

  14. Search for Le Sun in:

  15. Search for Lin Li in:

  16. Search for Wenwen Tang in:

  17. Search for Dianqing Wu in:

Contributions

D.W., L.L., Q.X., J.W., W.-J.W., W.T., M.S., and J.C. designed the experiments; Q.X., J.W., W.-J.W., S.C., Y.Z., M.S., W.T., and K.M. performed the experiments; D.W., L.L., Q.X., W.-J.W., M.S., J.W., A.L.M.B., L.C., and W.T. analyzed the data; X.Y. performed statistic and bioinformatic analyses; M.B., V.S., and L.S. created and provided important reagents; D.W., L.L., Q.X., J.W., W.-J.W., and W.T. wrote the manuscript; and all authors reviewed and approved the manuscript.

Competing interests

D.W. received research support from Just Biotherapeutic Asia, which licensed the intellectual property from Yale University on the basis of the findings reported in this article.

Corresponding authors

Correspondence to Lin Li or Wenwen Tang or Dianqing Wu.

Supplementary information

PDF files

  1. 1.

    Supplementary Figures & Tables

    Supplementary Figures 1–14 & Supplementary Tables 1–2

  2. 2.

    Life Sciences Reporting Summary