Cancer Immunology, Immunotherapy

, Volume 62, Issue 7, pp 1149–1159 | Cite as

Activation, dysfunction and retention of T cells in vaccine sites after injection of incomplete Freund’s adjuvant, with or without peptide

  • Elise P. Salerno
  • Sofia M. Shea
  • Walter C. Olson
  • Gina R. Petroni
  • Mark E. Smolkin
  • Chantel McSkimming
  • Kimberly A. Chianese-Bullock
  • Craig L. SlingluffJr.
Original Article


We conducted a randomized clinical trial in 45 patients with resected AJCC stage IIB-IV melanoma to characterize cellular and molecular events at sites of immunization with incomplete Freund’s adjuvant (IFA) alone, or a melanoma vaccine in IFA. At a primary vaccine site, all patients received a multi-peptide melanoma vaccine in IFA. At a replicate vaccine site, which was biopsied, group 1 received IFA only; group 2 received vaccine in IFA. Lymphocytes isolated from replicate vaccine site microenvironments (VSME) were compared to time-matched peripheral blood mononuclear cells (PBMC) in ELISpot and flow cytometry assays. Compared to PBMC, the VSME had fewer naïve and greater proportions of effector memory CD8+ T cells (TCD8). The vast majority of TCD8 within the VSME were activated (CD69+), with a concentration of antigen-specific (tetramerpos) cells in the VSME, particularly in vaccine sites with peptide (group 2). CXCR3+ lymphocytes were concentrated in the VSME of all patients, suggesting IFA-induced chemokine recruitment. TCD8 expression of retention integrins αEβ7 and α1β1 was elevated in VSME, with the highest levels observed in antigen-specific cells in VSME containing peptide (group 2). TCD8 retained in the VSME of both groups were strikingly dysfunctional, with minimal IFN-γ production in response to peptide stimulation and few tetramerpos cells producing IFN-γ. These data suggest that vaccine-induced selective retention and dysfunction of antigen-specific TCD8 within VSME may represent a significant mechanism underlying transient immune responses and low clinical response rates to peptide vaccines administered in IFA.


Human  T cell  Incomplete Freund’s adjuvant  Peptide vaccine  Melanoma  CIMT 2012 Integrins Melanoma vaccine 



Cutaneous leukocyte antigen


High endothelial venule


Incomplete Freund’s adjuvant


Peripheral blood mononuclear cell


Central memory T cell (CD45RAneg/CCR7+)


Effector memory T cell (CD45RAneg/CCR7neg)


CD45RApos effector memory T cell (CD45RAneg/CCR7neg)


Tertiary lymphoid organ


Naïve T cell (CD45RA+/CCR7+)


Vaccine site microenvironment



We greatly appreciate the work of Patrice Neese, Carmel Nail, and Priscilla Merrill in care of the patients on this trial, and Cheryl F.Murphy, Nadejda V. Galeassi, Kelly Smith and Elizabeth Coleman in immune analyses and data management. Funding support was provided by the National Institutes of Health Research Training Grant T32HL007849 (PI- Irving Kron) to EPS and the University of Virginia Rebecca Clay Harris Memorial Fellowship to SMS. Additional funding for this study was provided by the University of Virginia Cancer Center Support Grant (NIH/NCI P30 CA44579, Biorepository and Tissue Research Facility) and the University of Virginia General Clinical Research Center (NIH M01 RR00847). Peptides used in this vaccine were prepared with philanthropic support from the Commonwealth Foundation for Cancer Research and Alice and Bill Goodwin. Additional philanthropic support was provided by the James and Rebecca Craig Foundation, George S. Suddock, Richard and Sherry Sharp, and the Patients and Friends Research Funds of the University of Virginia Cancer Center. Montanide ISA-51 (produced by Seppic, Inc.) was used in the vaccines of this trial, but paid for by the University of Virginia. No corporate funding support was provided for this study.

Conflict of interest

The authors declare that they have no conflict of interest.

Supplementary material

262_2013_1435_MOESM1_ESM.pdf (391 kb)
Supplementary material 1 (PDF 390 kb)


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Copyright information

© Springer-Verlag Berlin Heidelberg 2013

Authors and Affiliations

  • Elise P. Salerno
    • 1
  • Sofia M. Shea
    • 2
  • Walter C. Olson
    • 1
  • Gina R. Petroni
    • 3
  • Mark E. Smolkin
    • 3
  • Chantel McSkimming
    • 4
  • Kimberly A. Chianese-Bullock
    • 1
  • Craig L. SlingluffJr.
    • 1
    • 5
  1. 1.Division of Surgical Oncology, Department of SurgeryUniversity of VirginiaCharlottesvilleUSA
  2. 2.Department of PathologyUniversity of VirginiaCharlottesvilleUSA
  3. 3.Department of Public Health SciencesUniversity of VirginiaCharlottesvilleUSA
  4. 4.Cardiovascular Research CenterUniversity of VirginiaCharlottesvilleUSA
  5. 5.UVA Health SystemCharlottesvilleUSA

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