561 / 1096
S546
ESTRO 36
_______________________________________________________________________________________________
4
Candiolo Cancer Institute- Torino- Italy, Medical Physics
Unit, Candiolo TO, Italy
Purpose or Objective
The important goal of tumor immunotherapy is to identify
strategies to modulate
in vivo
the
anti-tumor immunity in
order to achieve clinical efficacy. In the last decade great
progresses have been made in the field of tumor
immunology and an increasing number of tumor antigens
have been identified. In particular, several different
engineering T-cells were designed to express receptors
specific for antigens expressed in the tumor compartment.
Herein, in order to block tumor progression, we employed
Chimeric Antigen Receptors (CARs) technology to target
the tumor vasculature. To this aim we used a spontaneous
mouse tumor model of pancreatic neuroendocrine
insulinoma, RIP-Tag2. Despite its low frequency in cancer
patients, we chose this model, since develops invasive
tumors through well-characterized and synchronous pre-
malignant stages, sustained by active angiogenesis. Little
is known about the effects of ionizing radiation on tumor
burden of RIP-Tag2 mice and the impact of whole-body
irradiation on the overall survival.
Material and Methods
In the current study tumor-bearing RIP-Tag2 mice were
irradiated with two different sub-lethal dosages (5 and 6
Gy respectively), by means of 6MV x rays of Tomotherapy;
a treatment plan was performed and evaluated for each
cage pie . We hypothesized that sub-lethal radiation might
be more effective than a lethal dose radiation and
clinically acceptable in promoting anti-tumor immunity.
The day after the irradiation mice were injected with
enginered T-cells (20 million cells).
Results
Firstly,we observed that irradiation
per se
did not affect
tumor growth and all the mice survived until the end of
treatment. Furthermore, by means of this approach, we
found statistically significant inhibition of tumor growth in
mice treated with anti-vasular CAR-T-cells compared with
controls. Interestingly, we noticed a strong reduction in
tumor vessel area and a decrease of blood vessel
permeability.
Conclusion
These data suggest that the tumor vasculature can be
efficiently targeted by specific CAR-T-cells, causing a
significant reduction in the tumor burden of RIP-Tag2 mice
and potentially in other tumor types.
PO-0990 Combining radiotherapy and notch inhibition
in melanoma
K. Thippu Jayaprakash
1,2
, M. Hussein
3
, A. Nisbet
3,4
, R.
Shaffer
2
, M. Ajaz
1,2
1
University of Surrey, Department of Clinical and
Experimental Medicine, Guildford, United Kingdom
2
St Luke's Cancer Centre- Royal Surrey County Hospital,
Department of Oncology, Guildford, United Kingdom
3
St Luke's Cancer Centre- Royal Surrey County Hospital,
Department of Medical Physics, Guildford, United
Kingdom
4
University of Surrey, Department of Physics, Guildford,
United Kingdom
Purpose or Objective
Melanoma is classically viewed as a radioresistant tumour.
Phenotypic plasticity, specifically the emergence of a
cancer stem cell (CSC) population, may be one reason for
this. The notch signalling pathway plays a crucial role in
maintenance of the CSC phenotype, and also in cell
migration. This pathway is frequently aberrant in
melanoma and is therefore a potential mechanism for the
observed radioresistance. The aims of this project were
(1) to investigate whether notch inhibition with the γ -
secretase inhibitor, RO4929097 that targets γ - secretase
cleavage and thereby inhibiting the notch signalling
pathway, improves the radiation sensitivity of melanoma
cell lines; (2) to investigate the effects of notch inhibition
and radiotherapy on melanoma cell migration.
Material and Methods
Two melanoma cell lines, A375 and SKMEL28 were
irradiated with 250 kV x-rays to 1, 2, 4, 6, 8 and16 Gy in
combination with 1, 3, 10, 30 and 100 µm of RO4929097 in
96-well plates. Cells were permitted to grow for a further
5 - 7 days and viability was assessed with the MTS assay.
Loewe’s combination index (CI) was used (CI=(C
A
,
X
/IC
X,
A
)+(C
B
,
X
/IC
X, B
)) to evaluate the interaction between
radiation and RO4929097. For cell migration experiments,
A375 and SKMEL28 cells were treated with 10 and 100 µm
of RO4929097, alone and in combination with radiation (2
and 8 Gy) in 6-well plates. A scratch was performed and
daily light field microscope photographs were taken. In all
experiments, radiation was delivered one hour after cells
were treated with RO4929097.
Results
Loewe’s CI of < 1 and > 1 are taken to indicate synergism
and antagonism respectively. The Loewe’s combination
index analysis reproducibly showed strong synergy in A375
melanoma cells when radiation doses of 1, 2, 4, 6 and 8
Gy were combined with 100 µm of RO4929097 and a trend
towards mild synergy was observed with lower doses of
radiation and higher doses of RO4929097
(Figure 1
). This
may be due to a reduction in the number of CSCs by
RO4929097 that renders lower radiation doses more
effective. Similar patterns of interaction were observed
for SKMEL28 cells.
Cell migration assays showed that cell migration was
inhibited in both cell lines following treatment with 10 and
100 µm of RO4929097 and this was more pronounced at
100 µm, and similar effects were seen when radiation was
combined with RO4929097. 8 Gy alone failed to control
cellular migration but this was abrogated by the addition
of RO4929097
(Figure 2)
.
Conclusion
Inhibition of the notch signalling pathway increases the
radiosensitivity of melanoma cells. We hypothesise this is