Blood Journal
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Small molecule Toll-like receptor 7 agonists localize to the MHC class II loading compartment of human plasmacytoid dendritic cells

  1. Carla Russo1,
  2. Ivan Cornella-Taracido2,
  3. Luisa Galli-Stampino1,
  4. Rishi Jain2,
  5. Edmund Harrington2,
  6. Yuko Isome2,
  7. Simona Tavarini1,
  8. Chiara Sammicheli1,
  9. Sandra Nuti1,
  10. M. Lamine Mbow3,
  11. Nicholas M. Valiante3,
  12. John Tallarico2,
  13. Ennio De Gregorio1, and
  14. Elisabetta Soldaini1
  1. 1Novartis Vaccines & Diagnostics, Siena, Italy;
  2. 2Novartis Institutes for Biomedical Research, Cambridge, MA; and
  3. 3Novartis Vaccines & Diagnostics, Cambridge, MA
  1. Presented in abstract form at the DC2010: Forum on Vaccine Science, Lugano, Switzerland, September 26-30, 2010.

Abstract

TLR7 and TLR8 are intracellular sensors activated by single-stranded RNA species generated during viral infections. Various synthetic small molecules can also activate TLR7 or TLR8 or both through an unknown mechanism. Notably, direct interaction between small molecules and TLR7 or TLR8 has never been shown. To shed light on how small molecule agonists target TLRs, we labeled 2 imidazoquinolines, resiquimod and imiquimod, and one adenine-based compound, SM360320, with 2 different fluorophores [5(6) carboxytetramethylrhodamine and Alexa Fluor 488] and monitored their intracellular localization in human plasmacytoid dendritic cells (pDCs). All fluorescent compounds induced the production of IFN-α, TNF-α, and IL-6 and the up-regulation of CD80 and CD86 by pDCs showing they retained TLR7-stimulating activity. Confocal imaging of pDCs showed that, similar to CpG-B, all compounds concentrated in the MHC class II loading compartment (MIIC), identified as lysosome-associated membrane protein 1+, CD63, and HLA-DR+ endosomes. Treatment of pDCs with bafilomycin A, an antagonist of the vacuolar-type proton ATPase controlling endosomal acidification, prevented the accumulation of small molecule TLR7 agonists, but not of CpG-B, in the MIIC. These results indicate that a pH-driven concentration of small molecule TLR7 agonists in the MIIC is required for pDC activation.

  • Submitted December 29, 2010.
  • Accepted March 28, 2011.
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