Supplementary Materials

Supplementary Material for:

In vivo imaging reveals a tumor-associated macrophage–mediated resistance pathway in anti–PD-1 therapy

Sean P. Arlauckas, Christopher S. Garris, Rainer H. Kohler, Maya Kitaoka, Michael F. Cuccarese, Katherine S. Yang, Miles A. Miller, Jonathan C. Carlson, Gordon J. Freeman, Robert M. Anthony, Ralph Weissleder, Mikael J. Pittet*

*Corresponding author. Email: mpittet{at}mgh.harvard.edu

Published 10 May 2017, Sci. Transl. Med. 9, eaal3604 (2017)
DOI: 10.1126/scitranslmed.aal3604

This PDF file includes:

  • Materials and Methods
  • Fig. S1. AF647–aPD-1 mAb quantification in various tissues at 24 hours after injection.
  • Fig. S2. Specificity of T cell reporter mice and dextran nanoparticles.
  • Fig. S3. Representative intravital microscopy time course of AF647–aPD-1 from an IFN-γ–YFP mouse.
  • Fig. S4. T cell and macrophage motility before and after aPD-1 treatment.
  • Fig. S5. Assessment of aPD-1 binding to tumor cells.
  • Fig. S6. aPD-1 transfer to tumor macrophages in vivo.
  • Fig. S7. Distribution of AF647–aPD-1 across tumor models.
  • Fig. S8. PD-1 expression by CD8+ T cells in the MC38 tumor microenvironment.
  • Fig. S9. aPD-1 mAb transfer from T cells to macrophages in vitro.
  • Fig. S10. Assessment of aPD-1 internalization after binding to PD-1.
  • Fig. S11. aPD-1 degradation by macrophages.
  • Fig. S12. PD-1 localization following aPD-1 internalization after binding to PD-1.
  • Fig. S13. Comparative analysis of mAb glycosylation patterns between mouse and human PD-1 antibodies.
  • Fig. S14. Assessment of cell membrane component exchange during aPD-1 transfer.
  • Fig. S15. Confirmation of deglycosylation and antigen binding affinity for rat anti-mouse PD-1.
  • Fig. S16. Impact of Fc blockade on aPD-1 treatment efficacy.
  • Fig. S17. Proposed resistance mechanism and potential improvement of aPD-1 mAb therapy.
  • Legends for movies S1 to S4
  • References (4652)

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Other Supplementary Material for this manuscript includes the following:

  • Movie S1 (.mov format). Intravital microscopy imaging of AF647–aPD-1 injection in a DPE-GFP mouse bearing an MC38–H2B-mApple tumor in a dorsal skinfold chamber.
  • Movie S2 (.mp4 format). Intravital microscopy imaging of AF647–aPD-1 injection in an IFN-γ–enhanced YFP GREAT mouse bearing an MC38–H2B-mApple tumor in a dorsal skinfold chamber.
  • Movie S3 (.mov format). Live imaging of mouse T cell aPD-1 transfer to macrophages.
  • Movie S4 (.mov format). Live imaging of human T cell aPD-1 transfer to macrophages.