IMMUNOTHERAPY AGAINST THE RADIAL GLIA MARKER GLAST EFFECTIVELY TRIGGERS SPECIFIC ANTITUMOR EFFECTORS WITHOUT AUTOIMMUNITY
Creators: name: CANTINI, GABRIELE
Publisher: name: Universit� degli Studi di Milano
Year: 2014

description: Glioblastoma (GB) is the most aggressive and fatal tumor of the central
nervous system (CNS). Despite promising results of different therapeutic
approaches, conventional treatments such as surgery, radiation and
chemotherapy with Temozolomide (TMZ) are largely palliative. The
identification of antigens preferentially expressed in GB and involved in its
malignant phenotype is critical for developing therapeutic strategies.
Murine and human GB contain a fraction of cancer cells with stem-like
features (CSC) and it has been proposed that only this population may be
responsible for glioma recurrence. We previously found that GLAST, a
membrane protein with relevant action in glutamate trafficking, is highly
expressed in murine GL261 glioma stem-like cells (GSC).
In this study, we first investigated the functional consequences of GLAST
enrichment using an immune-magnetic sorting approach to isolate GLAST+
cells from murine GSC. GLAST+ cells were significantly more tumorigenic
than GLAST- or unsorted cells (p&lt; 0.0006) when injected intracranially,
showing the highly invasive nature of this subpopulation. GLAST
expression and its impact on aggressiveness in vivo constituted a
background of translational relevance.
To determine whether GLAST peptide-based immunotherapy protects
against GL261-gliomas, we treated glioma-bearing mice with three
subcutaneous injections of four GLAST-derived peptides emulsified with
Montanide? ISA 51 VG in association with granulocyte macrophage
colony-stimulating factor (GM-CSF) injections. We found that immunization
with GLAST peptides efficiently induces specific anti-tumor response in the
murine GL261-glioma preventing the tumor progression in 40% of
immunized mice. The immunization induced relevant effects on tumor
microenvironment by up-regulating IFN? and TNF? as well as downregulating
TGF?1 and ?2. GLAST expression significantly decreased in
III
gliomas from immunized mice, as evaluated by histological analysis and
real-time PCR. Beyond these changes, trafficking of NK cells, CD8+ T cells
and CD4+ T cells in the spleens and lymph nodes and their homing ability
into the brain significantly increased in immunized mice when compared
with controls.
A local chemotactic gradient characterized by expression of CXCL10
(which may be responsible for the recruitment of CTL), CCL3, CCL4 and
CCL5 (which are involved in NK cell migration), and NKG2D ligand in
immunized mice may play a role in the accumulation of immune cells
(particularly of NK cells) at the sites of tumor formation.
Autoimmune reactions were not observed in immunized mice supporting
evidence that GLAST may constitute a glioma antigen against which
immune responses can be efficiently induced without toxicity.
In order to emphasize the translational relevance of GLAST as marker and
glioma-associated antigen we will extend our studies to human GB.
GLAST expression was investigated by immunohistochemistry in 39 human
primary GB. High percentage of GLAST+ cells, and a moderate or strong
reactivity, was associated with a decreased overall survival (p= 0.02).
The main result of this study is the definition of GLAST as an attractive
glioma-associated antigen for targeting in peptide-based immunotherapy in
GB patients and a potential clinical marker in association with the patient
prognosis.