Abstracts for the 25th Annual Scientific Meeting of the International ...

J Immunother Volume 33, Number 8, October 2010
Abstracts
Histone Modification Upregulates Expression of IL-13Ra2
in Human Pancreatic Cancer Cells and Enhances the
Effectiveness of IL-13 Receptor Targeted Immunotoxin
IL-13-PE in Animal Model of Human Pancreatic Cancer
Toshio Fujisawa, Bharat H. Joshi, Raj K. Puri. Division of Cellular
Gene Therapy, Center for Biologics Evaluation & Research/FDA,
Bethesda, MD.
Interleukin 13 Receptor a2 (IL-13Ra2) is a cancer-associated
antigen and a target for receptor directed anti-cancer therapy. We
have reported that a recombinant immunotoxin, consisting of
IL-13 and truncated Pseudomonas exotoxin (IL-13-PE) is highly
cytotoxic to tumors that express high density IL-13Ra2 in vitro and
in vivo. However, IL-13-PE is not very effective in tumors that
express low level of IL-13Ra2. Herein, we have examined whether
IL-13Ra2 can be modulated by epigenetic histone modification in
human pancreatic cancer cells. When eleven pancreatic cancer and
three normal (two epithelial and one fibroblast) cell lines were
treated by three different histone deacetylase (HDAC) inhibitors
(trichostatin A, sodium butyrate and Suberoylanilide hydroxamic
acid), we found that HDAC inhibitors significantly increased
mRNA and protein levels of IL-13Ra2 in low-IL-13Ra2 expressing
pancreatic cancer cells, compared to no or marginal upregulation
in high-IL-13Ra2 expressing pancreatic cancer and normal cells.
HDAC inhibitors enhanced anti-cancer effect of IL-13-PE in vitro
in low-IL-13Ra2 expressing pancreatic cancer cell lines by lowering
IC50 (concentration of IL-13-PE that kills 50% of the cells in
protein synthesis inhibition assay) from >1000 ng/mL to 40 to
50 ng/mL. Similar effects did not occur in high-IL-13Ra2 expressing
tumors or in normal human cell lines. In xenograft mouse
models developed by implanting low-IL-13Ra2 expressing cell lines
(Panc-1 and ASPC-1), IL-13-PE alone showed no anti-cancer
effect, but in combination with HDAC inhibitors (trichostatin A
and Suberoylanilide hydroxamic acid), IL-13-PE significantly
decreased tumor burden and prolonged survival of animals.
Trichostatin A increased IL-13Ra2 expression in tumors approximately
500 times compared to vehicle treated mice with Panc-1
tumors. A combination therapy with HDAC inhibitors and IL-13-
PE had marginal or no detectable histological changes in any vital
organs of mice, which indicate that this effect is tumor specific
restricted within tumor compartment. Taken together, we have
identified a new approach to target pancreatic cancer by combining
HDAC inhibitors with a potent immunotoxin IL-13-PE. Similar
approach may be useful for other malignancies as well.
COX2 Blockade Suppresses Gliomagenesis by Inhibiting
CCL2-Mediated Accumulation of Myeloid-derived Suppressor
Cells in Glioma Sites
Mitsugu Fujita*, Gary Kohanbash*, Heather A. McDonald*,
Louis Delamarre*, Stacy A. Deckerw, John R. Ohlfestw, Hideho
Okada*. *Neurological Surgery, University of Pittsburgh, Pittsburgh,
PA; w Pediatrics, University of Minnesota, Minneapolis, MN.
A line of epidemiologcal studies has suggested that the regular use
of nonsteroidal anti-inflammatory drugs (NSAIDs) is associated
with reduced glioma occurrence in humans. NSAIDs mediate their
biological effects at least partially by suppressing cyclooxygenase
(COX) 2 and its product prostaglandin (PG) E 2 , which induces
immunoregulatory cells including myeloid-derived suppressor cells
(MDSCs). PGE 2 also impacts chemokines, such as down-regulation
of CXCL10 production by dendritic cells. Based on these
findings, we hypothesized that COX2 blockade by NSAIDs would
suppress gliomagenesis by inhibiting MDSC accumulation in
gliomas. We chose acetylsalicylic acid (ASA) for immunological
evaluation because it did not inhibit the growth of cultured glioma
cells in vitro at pharmacological concentrations in contrast to
selective COX2 inhibitors. To address the impact of COX2
blockade on gliomagenesis, we induced de novo gliomas in mice
by trasposon-mediated intracerebroventricular transfection of
oncogenes. Daily ASA treatment inhibited systemic PGE 2 production
in wild type (WT) mice with developing gliomas. The
treatment prolonged the survival of these mice only when started
simultaneously with glioma induction. Glioma tissues of the ASAtreated
mice exhibited a decrease in mRNA for the MDSCattracting
chemokine Ccl2 but an increase of the CTL-attracting
chemokine Cxcl10. Consistently, analyses of brain tumor-infiltrating
leukocytes in the ASA-treated mice revealed a decrease of
CD11b + Ly6G + cells but an increase of CD8 + CD107a + cells
compared with those in non-treated mice. Cox2-deficient mice with
developing gliomas corroborated these phenomena. Ccl2-deficient
mice with developing gliomas exhibited a prolongation of survival
with a decrease in intratumoral MDSC accumulation. In addition,
these mice exhibited an increase in Cxcl10 and accumulation of
CD8+ cells in glioma sites. Antibody-mediated MDSC depletion
for WT mice with developing gliomas exhibited an increase in
Cxcl10 and prolonged survival, suggesting an impact of MDSCs on
glioma development. Cxcl10-deficient mice exhibited shortened
survival with a decrease in CD8+ cell accumulation, suggesting the
significance of CXCL10 in intratumoral T cell accumulation.
Collectively, these findings indicate important roles of COX2
pathway in gliomagenesis through CCL2-mediated MDSC accumulation
and reduction of CXCL10-mediated CTL accumulation
in glioma sites. These data also support development of NSAIDincluding
strategies for glioma immunotherapy.
Elevated MDSCs in Pancreatic and Esophago-gastric
Cancer Patients Correlates with Poor Prognosis
Nicola E. Annels*, Rachel F. Gabitass*w, Hardev Pandha*, Gary
W. Middletonw. *Postgraduate Medical School, University of
Surrey; w St Luke’s Cancer Centre, Royal Surrey County Hospital,
Guildford, United Kingdom.
Background: Myeloid derived suppressor cell (MDSC) and
regulatory T cell (Treg) accumulation in cancer patients is
proposed as an important mechanism of tumor immune evasion.
We present for the first-time analysis of both MDSCs and Tregs in
pancreatic and esophago-gastric cancer.
Methods: Peripheral blood samples were collected from 35
pancreatic, 46 esophageal and 19 gastric cancer patients and 31
normal healthy volunteers. PBMC was harvested and stored in
liquid N2 with subsequent flow cytometric analysis of MDSC
(HLADR- Lin1- CD33 + CD11b + ) and Treg (CD4+ CD25+
CD127low/- FoxP3+). Unpaired t tests were applied to analyse
MDSC and Treg levels between groups. Kaplan-Meier survival
analysis was performed to compare patients with normal MDSC
levels (as defined by the controls range, 0% to 2% total PBMCs)
and high MDSCs (>2%).
Results: The mean percentage of MDSCs in each cancer group was
significantly higher than controls; pancreatic 2.455% (P = 0.0005),
esophageal 1.623% (P = 0.0260) and gastric 2.029% (P = 0.0326),
with controls 1.104%. Pancreatic and esophago-gastric cancer
patients had significantly higher mean Treg percentages than
controls, P = 0.0036 and P = 0.0189 respectively. There was a
significant positive correlation between upper GI cancer patients’
MDSC and Treg scores (Spearman’s rho +0.285; P = 0.008).
Patients with normal MDSC levels survived longer than those with
high levels (log rank: P = 0.011), median survival 231 versus 145
days. The influence of MDSC score was further quantified by Cox
regression analysis with MDSC score as a covariate: P