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Collagen-binding IL-12 enhances tumour inflammation and drives the complete remission of established immunologically cold mouse tumours

Nature Biomedical Engineering volume 4, pages 531–543 (2020)Cite this article

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Abstract

Checkpoint-inhibitor (CPI) immunotherapy has achieved remarkable clinical success, yet its efficacy in ‘immunologically cold’ tumours has been modest. Interleukin-12 (IL-12) is a powerful cytokine that activates the innate and adaptive arms of the immune system; however, the administration of IL-12 has been associated with immune-related adverse events. Here we show that, after intravenous administration of a collagen-binding domain fused to IL-12 (CBD–IL-12) in mice bearing aggressive mouse tumours, CBD–IL-12 accumulates in the tumour stroma due to exposed collagen in the disordered tumour vasculature. In comparison with the administration of unmodified IL-12, CBD–IL-12 induced sustained intratumoural levels of interferon-γ, substantially reduced its systemic levels as well as organ damage and provided superior anticancer efficacy, eliciting complete regression of CPI-unresponsive breast tumours. Furthermore, CBD–IL-12 potently synergized with CPI to eradicate large established melanomas, induced antigen-specific immunological memory and controlled tumour growth in a genetically engineered mouse model of melanoma. CBD–IL-12 may potentiate CPI immunotherapy for immunologically cold tumours.

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Fig. 1: CBD–IL-12 binds to collagen with high affinity while retaining bioactivity.
Fig. 2: CBD–IL-12 induces regression of B16F10 melanoma and EMT6 mammary carcinoma.
Fig. 3: CBD–IL-12 induces intratumoural inflammation by rapidly localizing to the tumour.
Fig. 4: CBD–IL-12 minimizes irAEs in tumour-bearing and non-tumour-bearing mice.
Fig. 5: CBD–IL-12 decreases metastatic tumour burden by triggering the activation of innate and adaptive compartments of the immune system in the pulmonary metastatic model of B16F10 melanoma.
Fig. 6: CBD–IL-12 synergizes with CPI and elicits a tumour-antigen-specific response.

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Data availability

The main data supporting the results in this study are provided within the Article and its Supplementary Information. All data generated in this study, including source data and the data used to make the figures, are available from Figshare at https://doi.org/10.6084/m9.figshare.11971371.

References

  1. Larkin, J. et al. Combined nivolumab and ipilimumab or monotherapy in untreated melanoma. N. Engl. J. Med. 373, 23–34 (2015).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  2. Sharma, P. & Allison, J. P. The future of immune checkpoint therapy. Science 348, 56–61 (2015).

    Article  CAS  PubMed  Google Scholar 

  3. Rosenberg, S. A., Yang, J. C. & Restifo, N. P. Cancer immunotherapy: moving beyond current vaccines. Nat. Med. 10, 909–915 (2004).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  4. Langrish, C. L. et al. IL-12 and IL-23: master regulators of innate and adaptive immunity. Immunol. Rev. 202, 96–105 (2004).

    Article  CAS  PubMed  Google Scholar 

  5. McHeyzer-Williams, M. G. et al. Antigen-specific immunity. Immunol. Res. 22, 223–236 (2000).

    Article  CAS  PubMed  Google Scholar 

  6. Fallarino, F., Ashikari, A., Boon, T. & Gajewski, T. F. Antigen-specific regression of established tumors induced by active immunization with irradiated IL-12- but not B7-1-transfected tumor cells. Int. Immunol. 9, 1259–1269 (1997).

    Article  CAS  PubMed  Google Scholar 

  7. Rogge, L. et al. Selective expression of an interleukin-12 receptor component by human T helper 1 cells. J. Exp. Med. 185, 825–831 (1997).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Tugues, S. et al. New insights into IL-12-mediated tumor suppression. Cell Death Differ. 22, 237–246 (2015).

    Article  CAS  PubMed  Google Scholar 

  9. Lasek, W., Zagozdzon, R. & Jakobisiak, M. Interleukin 12: still a promising candidate for tumor immunotherapy? Cancer Immunol. Immunother. 63, 419–435 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. Kerkar, S. P. et al. IL-12 triggers a programmatic change in dysfunctional myeloid-derived cells within mouse tumors. J. Clin. Investig. 121, 4746–4757 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. Momin, N. et al. Anchoring of intratumorally administered cytokines to collagen safely potentiates systemic cancer immunotherapy. Sci. Transl. Med. 11, eaaw2614 (2019).

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  12. Ishihara, J. et al. Targeted antibody and cytokine cancer immunotherapies through collagen affinity. Sci. Transl. Med. 11, eaau3259 (2019).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Bekaii-Saab, T. S. et al. A phase I trial of paclitaxel and trastuzumab in combination with interleukin-12 in patients with HER2/neu-expressing malignancies. Mol. Cancer Ther. 8, 2983–2991 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. Atkins, M. B. et al. Phase I evaluation of intravenous recombinant human interleukin 12 in patients with advanced malignancies. Clin. Cancer Res. 3, 409–417 (1997).

    CAS  PubMed  Google Scholar 

  15. Gollob, J. A. et al. Phase I trial of concurrent twice-weekly recombinant human interleukin-12 plus low-dose IL-2 in patients with melanoma or renal cell carcinoma. J. Clin. Oncol. 21, 2564–2573 (2003).

    Article  PubMed  CAS  Google Scholar 

  16. Mariathasan, S. et al. TGFβ attenuates tumour response to PD-L1 blockade by contributing to exclusion of T cells. Nature 554, 544–548 (2018).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. Zhang, Y., Li, N., Suh, H. & Irvine, D. J. Nanoparticle anchoring targets immune agonists to tumors enabling anti-cancer immunity without systemic toxicity. Nat. Commun. 9, 6 (2018).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  18. Joyce, J. A. & Fearon, D. T. T cell exclusion, immune privilege, and the tumor microenvironment. Science 348, 74–80 (2015).

    Article  CAS  PubMed  Google Scholar 

  19. Gollob, J. A. et al. Phase I trial of twice-weekly intravenous interleukin 12 in patients with metastatic renal cell cancer or malignant melanoma: ability to maintain IFN-γ induction is associated with clinical response. Clin. Cancer Res. 6, 1678–1692 (2000).

    CAS  PubMed  Google Scholar 

  20. Ayers, M. et al. IFN-γ-related mRNA profile predicts clinical response to PD-1 blockade. J. Clin. Invest. 127, 2930–2940 (2017).

    Article  PubMed  PubMed Central  Google Scholar 

  21. Spranger, S., Dai, D., Horton, B. & Gajewski, T. F. Tumor-residing Batf3 dendritic cells are required for effector T cell trafficking and adoptive T cell therapy. Cancer Cell 31, 711–723 (2017).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Wang, L. L. et al. CXC-chemokine-ligand-10 gene therapy efficiently inhibits the growth of cervical carcinoma on the basis of its anti-angiogenic and antiviral activity. Biotechnol. Appl. Biochem. 53, 209–216 (2009).

    PubMed  Google Scholar 

  23. Ghiringhelli, F. et al. Activation of the NLRP3 inflammasome in dendritic cells induces IL-1β-dependent adaptive immunity against tumors. Nat. Med. 15, 1170–1178 (2009).

    Article  CAS  PubMed  Google Scholar 

  24. Liu, F. & Whitton, J. L. Cutting edge: re-evaluating the in vivo cytokine responses of CD8+ T cells during primary and secondary viral infections. J. Immunol. 174, 5936–5940 (2005).

    Article  CAS  PubMed  Google Scholar 

  25. Ryffel, B. Interleukin-12: role of interferon-γ in IL-12 adverse effects. Clin. Immunol. Immunopathol. 83, 18–20 (1997).

    Article  CAS  PubMed  Google Scholar 

  26. Neri, D. Antibody-cytokine fusions: versatile products for the modulation of anticancer immunity. Cancer Immunol. Res. 7, 348–354 (2019).

    Article  PubMed  PubMed Central  Google Scholar 

  27. Zhou, F. Molecular mechanisms of IFN-γ to up-regulate MHC class I antigen processing and presentation. Int. Rev. Immunol. 28, 239–260 (2009).

    Article  CAS  PubMed  Google Scholar 

  28. Clatza, A., Bonifaz, L. C., Vignali, D. A. & Moreno, J. CD40-induced aggregation of MHC class II and CD80 on the cell surface leads to an early enhancement in antigen presentation. J. Immunol. 171, 6478–6487 (2003).

    Article  CAS  PubMed  Google Scholar 

  29. Ho, P. C. et al. Immune-based antitumor effects of BRAF inhibitors rely on signaling by CD40L and IFNγ. Cancer Res. 74, 3205–3217 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  30. Pan, D. et al. A major chromatin regulator determines resistance of tumor cells to T cell-mediated killing. Science 359, 770–775 (2018).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  31. Boross, P. et al. Anti-tumor activity of human IgG1 anti-gp75 TA99 mAb against B16F10 melanoma in human FcgammaRI transgenic mice. Immunol. Lett. 160, 151–157 (2014).

    Article  CAS  PubMed  Google Scholar 

  32. Castle, J. C. et al. Exploiting the mutanome for tumor vaccination. Cancer Res. 72, 1081–1091 (2012).

    Article  CAS  PubMed  Google Scholar 

  33. Dankort, D. et al. Braf V600E cooperates with Pten loss to induce metastatic melanoma. Nat. Genet. 41, 544–552 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  34. Ishihara, J. et al. Matrix-binding checkpoint immunotherapies enhance antitumor efficacy and reduce adverse events. Sci. Transl. Med. 9, eaan0401 (2017).

    Article  PubMed  CAS  Google Scholar 

  35. Gros, A. et al. PD-1 identifies the patient-specific CD8+ tumor-reactive repertoire infiltrating human tumors. J. Clin. Investig. 124, 2246–2259 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  36. Spranger, S., Bao, R. & Gajewski, T. F. Melanoma-intrinsic β-catenin signalling prevents anti-tumour immunity. Nature 523, 231–235 (2015).

    Article  CAS  PubMed  Google Scholar 

  37. Drake, C. G., Lipson, E. J. & Brahmer, J. R. Breathing new life into immunotherapy: review of melanoma, lung and kidney cancer. Nat. Rev. Clin. Oncol. 11, 24–37 (2014).

    Article  CAS  PubMed  Google Scholar 

  38. Moynihan, K. D. et al. Eradication of large established tumors in mice by combination immunotherapy that engages innate and adaptive immune responses. Nat. Med. 22, 1402–1410 (2016).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  39. Garris, C. S. et al. Successful anti-PD-1 cancer immunotherapy requires T cell-dendritic cell crosstalk involving the cytokines IFN-γ and IL-12. Immunity 49, 1148–1161 (2018).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  40. Coughlin, C. M. et al. Tumor cell responses to IFNγ affect tumorigenicity and response to IL-12 therapy and antiangiogenesis. Immunity 9, 25–34 (1998).

    Article  CAS  PubMed  Google Scholar 

  41. Nagy, J. A., Chang, S. H., Dvorak, A. M. & Dvorak, H. F. Why are tumour blood vessels abnormal and why is it important to know? Br. J. Cancer 100, 865–869 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  42. Leonard, J. P. et al. Effects of single-dose interleukin-12 exposure on interleukin-12-associated toxicity and interferon-γ production. Blood 90, 2541–2548 (1997).

    CAS  PubMed  Google Scholar 

  43. Grigorian, A. & O’Brien, C. B. Hepatotoxicity secondary to chemotherapy. J. Clin. Transl. Hepatol. 2, 95–102 (2014).

    PubMed  PubMed Central  Google Scholar 

  44. van Herpen, C. M. et al. Intratumoral recombinant human interleukin-12 administration in head and neck squamous cell carcinoma patients modifies locoregional lymph node architecture and induces natural killer cell infiltration in the primary tumor. Clin. Cancer Res. 11, 1899–1909 (2005).

    Article  PubMed  Google Scholar 

  45. Mahvi, D. M. et al. Intratumoral injection of IL-12 plasmid DNA–results of a phase I/IB clinical trial. Cancer Gene Ther. 14, 717–723 (2007).

    Article  CAS  PubMed  Google Scholar 

  46. Swartz, M. A. & Lund, A. W. Lymphatic and interstitial flow in the tumour microenvironment: linking mechanobiology with immunity. Nat. Rev. Cancer 12, 210–219 (2012).

    Article  CAS  PubMed  Google Scholar 

  47. Halin, C. et al. Enhancement of the antitumor activity of interleukin-12 by targeted delivery to neovasculature. Nat. Biotechnol. 20, 264–269 (2002).

    Article  CAS  PubMed  Google Scholar 

  48. Fallon, J. et al. The immunocytokine NHS-IL12 as a potential cancer therapeutic. Oncotarget 5, 1869–1884 (2014).

    Article  PubMed  PubMed Central  Google Scholar 

  49. Strauss, J. et al. First-in-human phase I trial of a tumor-targeted cytokine (NHS-IL12) in subjects with metastatic solid tumors. Clin. Cancer Res. 25, 99–109 (2019).

    Article  PubMed  Google Scholar 

  50. Hank, J. A. et al. Immunogenicity of the Hu14.18-IL2 immunocytokine molecule in adults with melanoma and children with neuroblastoma. Clin. Cancer Res. 15, 5923–5930 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  51. Williford, J. M. et al. Recruitment of CD103+ dendritic cells via tumor-targeted chemokine delivery enhances efficacy of checkpoint inhibitor immunotherapy. Sci. Adv. 5, eaay1357 (2019).

    Article  PubMed  PubMed Central  Google Scholar 

  52. Sasaki, K. et al. Engineered collagen-binding serum albumin as a drug conjugate carrier for cancer therapy. Sci. Adv. 5, eaaw6081 (2019).

    Article  PubMed  PubMed Central  Google Scholar 

  53. Ishihara, J. et al. Improving efficacy and safety of agonistic anti-CD40 antibody through extracellular matrix affinity. Mol. Cancer Ther. 17, 2399–2411 (2018).

    CAS  PubMed  Google Scholar 

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Acknowledgements

We thank T. Gajewski (University of Chicago) for sharing Tyr:creER+ LSL-BrafV600E Ptenfl/fl and Tyr:creER+ LSL-BrafV600E Ptenfl/fl Ctnnb1STA mice; A. Solanki for assistance with tail vein injections; S. Gomes for assistance with experiments; T. Li for assistance with histology analysis; staff at the Human Tissue Resource Center of the University of Chicago for assistance with organ sectioning; and D. Leclerc (University of Chicago Flow Cytometry Core) for assistance with Luminex assays.

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  1. These authors contributed equally: Aslan Mansurov, Jun Ishihara.

Authors and Affiliations

  1. Pritzker School of Molecular Engineering, University of Chicago, Chicago, IL, USA

    Aslan Mansurov, Jun Ishihara, Peyman Hosseinchi, Lambert Potin, Tiffany M. Marchell, Ako Ishihara, John-Michael Williford, Aaron T. Alpar, Michal M. Raczy, Laura T. Gray, Melody A. Swartz & Jeffrey A. Hubbell

  2. Committee on Immunology, University of Chicago, Chicago, IL, USA

    Tiffany M. Marchell, Melody A. Swartz & Jeffrey A. Hubbell

  3. Ben May Department for Cancer Research, University of Chicago, Chicago, IL, USA

    Melody A. Swartz

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Contributions

A.M., J.I., M.A.S. and J.A.H. designed the experiments and wrote the manuscript. A.M. and J.I. performed the experiments. P.H. assisted with the pulmonary metastasis model. L.P. assisted with the autochthonous melanoma model. T.M.M. assisted with the antigen restimulation experiment and prepared B16F10 exosomes. A.I. blindly evaluated histological sections. J.M.-W. and L.T.G. assisted with tumour experiments. A.T.A. assisted with blood chemistry analysis. M.M.R. assisted with and analysed MALDI-TOF data.

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Correspondence to Jun Ishihara or Jeffrey A. Hubbell.

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Competing interests

J.I., A.I., M.A.S. and J.A.H. are inventors on U.S. Provisional Patent applications 62/638,520, 28/984,351 and 62/727,156. J.I., A.I., M.A.S. and J.A.H. are founders and shareholders in Arrow Immune Inc., which is developing the technology presented in this report, and M.A.S. and J.A.H. have leadership roles in that company. The other authors declare no competing interests.

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Mansurov, A., Ishihara, J., Hosseinchi, P. et al. Collagen-binding IL-12 enhances tumour inflammation and drives the complete remission of established immunologically cold mouse tumours. Nat Biomed Eng 4, 531–543 (2020). https://doi.org/10.1038/s41551-020-0549-2

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