Abstract Book 2010 - CIMT Annual Meeting

078 Maccalli | Tumor biology & interaction with the immune system
Definition of the immunological properties of cancer stem cells
isolated from human glioblastoma
Cristina Maccalli 1 , Tiziano Di Tomaso 1 , Ena Wang 2 , Stefania Mazzoleni 3 , Gloria Sovena 1 ,
Soldano Ferrone 4 , Rossella Galli 3 , Francesco M. Marincola 2 , Giorgio Parmiani 1
1 Unit of Immuno-Biotherapy of Solid Tumors, San Raffaele Foundation Scientific Institute, Milan, Italy
2 Department of Tranfusion Medicine, National Institutes of Health, Bethesda, MD, USA
3 Neural Stem Cell Unit, San Raffaele Foundation Scientific Institute, Milan, Italy
4 Hillman Cancer Center, University of Pittsburgh Cancer Institute, Pittsburgh, USA
The main objectives of our study is to assess
whether cancer stem cells (CSCs) isolated from glioblastoma
multiforme (GBM) patients can be exploited
as source of antigens to elicit T cell-mediated
immune responses and their validation to design
immunotherapeutic protocols for GBM. We have
characterized, by immunofluorescence and cytofluorimetric
or confocal microscopy analysis, the
immune profile of 14 CSC and of 7 FBS tumor line
pairs isolated from GBM samples. Moreover, PBMCs
isolated from GBM patients were in vitro stimulated
with autologous CSCs and the specific reactivity of
T lymphocytes against GBM CSCs was evaluated by
IFN-g, Granzyme B release (ELISPOT) or CD107a
mobilization. We have assessed that both GBM CSC
and their FBS-cultured non-CSC pairs showed a
low immunogenic profile (MHC, NKG2DL and APM
molecules). Up-regulation of most of these molecules
was induced by IFNs or 5-Aza CdR, though
more efficiently in FBS than in CSCs. Notably, T
cell responses, both CD4+- and CD8+-mediated,
against GBM could be raised in one GBM patient by
stimulating in vitro PBMCs with autologous CSCs
pre-treated with IFN-β. However, TH2-mediated
responses with more efficient recognition of FBS
tumor cell than CSCs, could be found in 3 additional
GBM patients. One key finding of this study
is that CSCs but not FBS cells inhibit allogeneic T
cell proliferation. This immune-inhibitory activity
mediated by GBM CSCs was not dependent on the
secretion of TGFβ1 or 2, that were preferentially
down-modulated in CSC lines, nor on IL-10 and
IL1-3 that were undetectable in the supernatant of
these cell lines. Expression of the indoleamine 2,3dioxygenase
(IDO), a molecule implicated in the generation
of immune tolerance, was detected in both
CSCs and FBS tumor cell lines with high increase
following IFN-g treatment of the cells. Furthermore,
we found that both CSCs and FBS tumor cells
expressed immune response inhibitory molecules,
such as CTLA-4, PD-1 and PDL-1. Furthermore, a
differential gene signature that was confirmed at
the protein levels for some immunological-related
molecules was also found between CSC and FBS
lines. Moreover, this latter analysis provided information
of some candidate molecules associated
with CSCs that we are functionally investigating.
Altogether, these results indicate a lower immunogenicity
and higher suppressive activity of GBM
CSC compared to autologous FBS lines and the need
to identify immunomodulatory agents that can efficiently
restore the expression of immunogenic molecules
on CSCs.
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