Abstract Book 2010 - CIMT Annual Meeting

005 Vyth-Dreese | Therapeutic vaccination
MART-1 specific T cells after DNA tattoo vaccination of
melanoma patients
Florry Vyth-Dreese 1 , Martin van der Maas 1 , Willeke van de Kasteele 1 , Trees Dellemijn 1 ,
Sandra Adriaansz 2 , Henk Mallo 2 , Bastiaan Nuijen 3 , Christian Ottensmeier 4 , Lindsey Low 4 ,
Christian Blank 1,2 , Ton Schumacher 1 and John Haanen 1,2
1 Division of Immunology, The Netherlands Cancer Institute-Antoni van Leeuwenhoek Hospital,
1066 CX Amsterdam, The Netherlands
2 Medical Oncology Department, The Netherlands Cancer Institute-Antoni van Leeuwenhoek
Hospital, 1066 CX Amsterdam, The Netherlands
3 Pharmacy, Slotervaart Hospital, 1066 EC Amsterdam, The Netherlands
4 Cancer Sciences Division, University of Southampton, Southampton SO16 6YD, United Kingdom
Recently, we developed a novel, rapid and potent
intradermal DNA vaccination method, called DNA
tattooing. Previous studies in experimental systems
showed a strong increase in vaccine-specific T cells
compared to intramuscular vaccination, parallelled
by robust anti-tumor T cell responses and rejection
of established subcutaneous tumors. These
results prompted us to initiate a phase I clinical
study with an inhouse manufactured GMP-grade
DNA vaccine, pDermatt. Here we report data from
the immunomonitoring of our first phase I clinical
trial of DNA tattoo vaccination of stage IV melanoma
patients using this DNA vaccine encoding a
tetanus toxin fragment c-modified MART-1 epitope
fusion protein.
Sofar, six patients were treated with DNA tattoo
vaccination on day 0, 3 and 6 as a prime and on day
28, 31 and 34 as a boost vaccination. Per cohort of
3 patients the vaccine dose was doubled from 0.5
mg to 1 mg DNA per tattoo. Flow cytometry and
EliSpot assays were performed to examine the presence
and specificity of T cells in peripheral blood
samples, taken before therapy and at week 2, 4, 6
and 8. Biopsies were taken from the vaccination site
at week 2 and 6 and examined for the presence of
immune cell subsets and specific T cells by immunohistochemistry
and flow cytometry.
To enable flow cytometric analysis for specific T
cells, biopsies were taken from the vaccination
site and cultured in vitro for 2 weeks in high dose
IL-2 (6000 IU/ml). In 5/6 patients, compared to 0/6
normal skin biopsies obtained from breast cancer
operations, this resulted in outgrowth of MART-
1-specific CD8 T lymphocytes, with 3/5 patient
samples showing 30-50% MART-1 specific CD8 T
cells. Highest numbers were obtained from week
6 biopsies with a preferential increase in MART-1
modified specific T cells.
Virtually no therapy induced increases were observed
in percentages of MART-1 specific peripheral
blood CD8 T cells. EliSpot analysis revealed therapy
induced IFNγ responses to MART-1 modified and
wild type peptides in 2/3 patients tested in cohort
1. These responses followed similar, although
delayed, kinetics compared to tetanus fragment c
responses which served as internal controls.
Examination of immune cells at patient vaccination
sites showed similar numbers of CD8 T cells, compared
to normal skin, located in the subepidermal
or dermal, but not epidermal areas. An increase
was found for activated DC expressing CD80. From
these data it is concluded that DNA tattoo vaccination
induced MART-1 specific CD8 T cell migration
to the vaccination site.
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