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RT-PCR. Novel signalling interactions were characterised
by western blotting, and a series of cell-based assays.
Results
Exposure to fractionated 2Gy-irradition to a cumulative
total dose of 60Gy selected for 22Rv1 cells increase in
clonogenic survival following irradiation (1.3 fold increase
in survival after 2Gy and 2.2 fold increase after 10Gy)
when compared to both parent 22Rv1 and aged-matched
controls. The cross-evaluation strategy of the molecular
modifications associated with a combination of
radiobiological factors has identified miR-4284 as down
regulated amongst radiation resistant models. Further
evaluation of this miRNA indicates interaction with RLIM,
RASGEF1, YB-1 and Notch-3. YB-1 inhibition with Fisetin
significantly reduced clonogenic survival following
irradiation, and modified Notch-3 receptor activation.
Analysis of RNA extracted from a series of pooled samples
from prostate cancer patients identified elevation of
notch-3 mRNA levels in higher grade and hypoxic tumours.
Validation in cell lines further identifies modification in
Notch-3 activation following 5-Gy irradiation.
Conclusion
This study identifies novel molecular radiobiology that
may explain the multiple effects of radiation on the
molecular biology of prostate cancer cells. This work has
the potential to influence future direction of suitability
and treatment of radiotherapy prostate patients.
PV-0372 Histology-specific quantitative mapping and
targeting of glucose and glutamine metabolism in
NSCLC
T.W.H. Meijer
1
, P.N. Span
1
, W.J.M. Peeters
1
, R.
Biemans
3
, L.F. De Geus-Oei
2
, D. Vriens
2
, L.J. Dubois
3
, J.
Bussink
1
1
UMC St Radboud Nijmegen, Radiation Oncology,
Nijmegen, The Netherlands
2
Leiden university medical center, Radiology, Leiden,
The Netherlands
3
Maastricht University Medical Centre, Radiation
Oncology, Maastricht, The Netherlands
Purpose or Objective
Increased glycolysis and glutamine use are related to
resistance to radiotherapy. Therefore, targeting tumor
cell metabolism may improve radiotherapy efficacy in
NSCLC. In this prospective cohort study, we describe
pharmacokinetic rate constants of
18
F-FDG metabolism
(K
1
-k
3
) and fractional blood volume (V
B
) in regions with
different levels of glucose metabolic rate (MR
glc
) and
compare these between the major NSCLC histological
subtypes (adeno- (AC) and squamous cell carcinomas
(SCC)). Furthermore, glycolytic rate and growth delay plus
apoptotic index by glucose and/or glutamine inhibition
were assessed in six NSCLC cell lines
in vitro.
Material and Methods
One-hour dynamic
18
F-FDG-PET/CTs were acquired in 38
NSCLC patients (tumor size at least 30 mm in diameter).
Parametric images of Patlak MR
glc
values were obtained.
Lesions were delineated using the fuzzy locally adapted
Bayesian (FLAB) algorithm. Tumors were divided into
three equal volumes of increasing MR
glc
, in which K
1
-k
3
and
V
B
were computed.
For
in vitro
experiments, AC (H522, HCC827, H1975) and
SCC (H520, H226, SW900) NSCLC cell lines were used.
Glycolytic rate of cell lines was assessed by the
percentage extracellular acidification rate (% ECAR) under
normoxia and physiologic amount of glucose (i.e. 1.5 mM)
using Seahorse. Growth delay and apoptosis analyses were
performed under normoxia and 1.5 mM glucose using
IncuCyte. To examine the effect of metabolic inhibition
on growth delay and apoptotic index, the glycolysis
inhibitor lonidamine and/or glutaminase inhibitor 968
were used.
Results
In SCC compared to AC, lesion MR
glc
and k
3
were
significantly higher and V
B
was significantly lower. AC
showed less heterogeneity relative to SCC in terms of
mean MR
glc
, k
3
and V
B
. In SCC, a significant higher value for
k
3
and lower value for V
B
was found in regions with higher
MR
glc
.
Percentage ECAR under normoxic conditions was higher in
AC than SCC cell lines, corresponding to the presence of
18
F-FDG metabolism in areas with high V
B
in AC on dynamic
PET. Differences between histological subtypes were less
obvious in the inhibitor experiments. All cell lines show
decreased growth rate by glycolysis inhibition using
lonidamine. The combination of lonidamine with the
glutaminase inhibitor 968 was detrimental for cell growth.
In all cell lines, except H520, apoptotic index increased
using the combination therapy.
Conclusion
Adeno NSCLC show glycolysis under better
perfused/oxygenated conditions (aerobic glycolysis)
in
vivo
and
in vitro
, while SCC NSCLC exhibit anaerobic
glycolysis (high glycolytic rate under poor
vascularization). Apart from glucose, glutamine usage is
critical for these tumors. All cell lines show a marked
growth delay with increased apoptosis upon metabolic
inhibition with both lonidamine and 968. Therefore,
inhibiting metabolism might be a general approach to
optimize treatment, especially in combination with
radiotherapy in NSCLC.
PV-0373 Epigenetic and metabolic reprogramming as a
target for prostate tumor radiosensitization
A. Dubrovska
1
, C. Peitzsch
1
, A. Tyutyunnykova
1
, M. Cojoc
1
1
OncoRay - Center for Radiation Research in Oncology,
Medical Faculty Dresden Carl Gustav Carus- TU Dresden,
Dresden, Germany
Purpose or Objective
Radiotherapy remains one of the main modalities to treat
solid cancers and is one of the mainstays of curative
prostate cancer treatment. Nevertheless, the risk of
recurrence after radiotherapy still remains substantial in
locally advanced disease. Tumor relapse after
radiotherapy is attributed to the population of cancer
stem cells (CSCs) which survived the treatment.
Therefore, analysis of the CSC populations might be an
important predictive tool of radiotherapy outcome and
individualized treatment selection. However, compelling
evidence suggests a high plasticity of CSCs imposed by
tumor treatment. This study is aiming to investigate the
interconnection of the glutamine metabolism and cancer
cell plasticity in the development of tumor radioresistance
for the development of new biomarkers to predict
radiation treatment outcome.
Material and Methods
The employed methodological approaches include gene
expression analysis, comparative genomic hybridization
array, proteomic analysis, metabolic profiling, in vitro
radiobiological clonogenic survival assays, assessment of
the histone methylation marks and CSC marker expression,
analysis of DNA damage repair and oxidative stress
response. This study is based on the different models
including tumor cell lines and their radioresistant
derivatives, prostate cancer xenografts, ex vivo treated
tissues and analysis of the publicly available TCGA
prostate cancer datasets.
Results
Our study revealed that irradiation causes long-term
upregulation in the expression of stem cell markers and
induces tumor cell reprogramming. Furthermore,
radioresistant and tumorigenic cell populations undergo a
phenotypic switch during the course of radiotherapy. This
phenotypic plasticity is associated with genetic,
epigenetic and metabolic changes induced by irradiation.
Expression of CSC markers and proteins involved in