The impact of γ-irradiation on the induction of bystander killing by genetically engineered ovarian tumor cells: implications for clinical use
Zweiri and Christmas Cancer Cell Int
The impact of γ-irradiation on the induction of bystander killing by genetically engineered ovarian tumor cells: implications for clinical use
Jehad Zweiri 0 1
Stephen E. Christmas 1
0 Dept. of Clinical Infection & Immunology, Institute of Infection & Global Health , Ronald Ross Building , University of Liverpool , 8, West Derby St, Liverpool L69 7BE , UK
1 Institute of Infection & Global Health, University of Liverpool , Liverpool , UK
Background: Cellular based therapeutic approaches for cancer rely on careful consideration of finding the optimal cell to execute the cellular goal of cancer treatment. Cell lines and primary cell cultures have been used in some studies to compare the in vitro and in vivo efficacy of autologous vs allogeneic tumour cell vaccines. Methods: This study examines the effect of γ-irradiation on a range of tumor cell lines in conjunction with suicide gene therapy of cancer. To determine the efficacy of this modality, a series of in vitro and in vivo experiments were conducted using genetically modified and unmodified tumor cell lines. Results: Following co-culture of HSV-TK modified tumor cells and unmodified tumor cells both in vitro and in vivo we observed that the PA-STK ovarian tumor cells were sensitive to γ-irradiation, completely abolishing their ability to induce bystander killing of unmodified tumor cells. In contrast, TK-modified human and mouse mesothelioma cells were found to retain their in vitro and in vivo bystander killing effect after γ-irradiation. Morphological evidence was consistent with the death of PA-STK cells being by pyknosis after γ-irradiation. These results suggest that PA-STK cells are not suitable for clinical application of suicide gene therapy of cancer, as lethal γ-irradiation (100 Gy) interferes with their bystander killing activity. However, the human mesothelioma cell line CRL-5830-TK retained its bystander killing potential after exposure to similarly lethal γ-irradiation (100 Gy). CRL-5830 may therefore be a suitable vehicle for HSVTK suicide gene therapy. Conclusions: This study highlights the diversity among tumor cell lines and the careful considerations needed to find the optimal tumor cell line for this type of suicide gene therapy of cancer.
Tumor cell lines; γ-irradiation; Suicide gene therapy; Anti-tumor immune response; Cell death; Bystander killing effect; Cancer clinical trials
The central objective in cancer therapy is to kill the
malignant cells while causing little or preferably no
collateral damage to healthy cells. Suicide gene therapy, as
applied to the treatment of cancer, holds the potential
to achieve just that [
]. An example is the insertion
of the herpes simplex virus thymidine kinase (HSV-TK)
gene into cancer cells which are consequently induced to
“commit suicide” when in the presence of otherwise
nontoxic doses of ganciclovir (GCV) [
]. This selective
toxic effect of the purine analogue ganciclovir is because
HSV-TK phosphorylates ganciclovir, converting it
ultimately to ganciclovir-triphosphate, a toxic compound
when inserted into the DNA of these transfected cells
]. As with any other gene therapy strategy, and any
anti-cancer treatment approach, its main limitation is
the selective targeting and transduction of all tumor cells
in vivo [
]. However, it may not be necessary to
transduce every tumor cell in vivo to bring about a
clinicallymeaningful anti-tumor effect [
]. Indeed, it has been
demonstrated that two types of “bystander tumor cell
killing” mechanisms are mediated by this approach: (a) a
“direct” bystander effect, due to the transfer of
ganciclovir triphosphate from HSV-TK-positive tumor cells into
untransfected neighboring cells [
], (b) a systemic
immunologically-mediated bystander effect due to the
in vivo immune presentation of
tumor-specific/associated antigens following the killing of
HSV-TK—expressing cells [
The genetically modified HSV-TK human ovarian
cancer cell line, PA1-STK, has been used for the
treatment of solid tumors (administered intraperitoneally in
patients with ovarian cancer) [
]. The in vitro culture
of these cells in the presence of ganciclovir provoked
bystander killing, but with limited cytotoxic activity
in vivo [
]. The rationale for this strategy was that
PASTK cells, injected in the vicinity of the patient’s tumor
bulk, could make contact with, and seed themselves onto,
the patient’s tumor cells in vivo and, after treatment with
ganciclovir, could commit suicide and kill the patient’s
tumor cells by a “direct” bystander mechanism (e.g. gap
junction mediated transfer of the phosphorylated
ganciclovir from the HSV-TK positive cells to the TK negative
cells. This direct cytotoxicity could then induce a more
systemic immunological bystander tumor-killing effect
Irradiation in conjunction with cancer gene therapy
may have several potential benefits. As well as synergistic
killing of tumor cells, it may enhance the bystander effect
by releasing tumor antigens, stimulating a broad
antitumor immune response [
]. However, it is
important to establish that irradiated HSV-TK-containing cells
are still able to induce direct bystander killing, without
contributing to the live tumor burden. This is a
particularly important consideration when such treatment is
considered for conditions of low tumor cell burden, but
poor prognosis, such as surgically debulked advanced
The objective of the present studies was to assess the
efficacy of bystander killing of unmodified tumour cells
by HSV-TK modified cells, before and after irradiation.
The studies reported here show that γ-irradiation has
a vastly different impact on the viability of tumor cells
in vitro; some die within hours whereas others remain
metabolically active for several days. The consequences
of such different sensitivities to γ-irradiation are that
the highly radiosensitive cells failed to mediate efficient
bystander killing either in vitro or in vivo, whilst the
less radiosensitive cells were able to mediate efficient
bystander killing both in vitro and in vivo. These studies
highlight the importance of sensitivity to γ-irradiation
for the development of allogeneic cell lines for the
clinical application of HSV-TK mediated bystander killing of
Human mesothelioma cell lines, CRL-5915, 5820, and
5830 were obtained from the American type culture
collection (Rockville, MD, USA) with the permission of Prof.
A Gazdar (MD Anderson Cancer Center, Texas, USA)
]. Mouse mesothelioma cell lines, ABI (H-2d) from
BALB/c mice, AE17 (H-2k) from CBA mice, and AC29
(H-2b) from C57BL/6 mice were obtained from Prof.
B. Robinson, QEII Medical Centre, University of
Western Australia, Nedlands, Australia [
]. Human ovarian
tumor (teratocarcinoma) cell line, PA-1 and PA-STK were
obtained from Prof. S Freeman, Tulane University
Medical School, New Orleans, USA [
]. Three ovarian tumor
cell lines already available in the laboratory were:
OVC432 (ovarian carcinoma, a kind gift from Dr C. Dohring,
Basel Institute for Immunology, Basel, Switzerland [
SKOV-3 (ovarian adenocarcinoma) [
], and OIB, an
ovarian tumor cell line established by Prof. F Farzaneh
in the Molecular Medicine Department [
]. All human
cell lines were maintained in DMEM, supplemented with
10% Fetal Calf Serum (FCS) and 1% sodium pyruvate. In
the case of mouse mesothelioma cells, the RPMI-medium
was supplemented with 5% FCS, 20 mM HEPES, 50 mM
2-mercaptoethanol (2ME) and 2 mM glutamine.
Cells to be infected were seeded at a density of 5 × 105
cells per 10 cm plate, 24 h before infection. Virus
producing cells were also grown to about 90% confluence, and
the medium changed 10 h prior to harvest of the culture
medium to ensure that fresh virus containing
supernatant was used [
The medium from virus producing cells was removed
and filtered through a 0.45 μm pore-size filter to remove
cell debris but allowing passage of the viral vector
through the filter. To enhance the retroviral infection,
8 μg/ml of polybrene (Sigma Aldrich, Poole, UK) was
added to the culture medium. Polybrene is required for
coating of target cells in order to neutralise their
negative surface charge, therefore increasing the efficiency
of infection [
]. The medium from the cells to be
infected was removed prior to infection, and the
vectorpolybrene mixture added. After 10 h of infection the
medium was changed. 48 h after infection, the medium
was removed and replaced with fresh medium containing
G418 at 1 mg/ml. G418 resistant clones were expanded.
In vitro GCV‑sensitivity studies on the tumor cell lines
The in vitro studies examining the effect of GCV on the
HSV-TK expressing mesothelioma and ovarian
carcinoma cells, as well as the untransduced cells, were
performed in 96-well plates. Transduced and untransduced
cells were plated in triplicate at two densities, 104 cells/
well and 105 cells/well for comparison. After 2 days, the
medium was replaced by fresh DMEM containing the
indicated concentrations of GCV in the range 10,000–
0.01 μM. Cells were then incubated at 37 °C in
humidified 5% CO2 for 5 days. Sensitivity to GCV treatment
was measured by using a colorimetric cell proliferation
assay that measures viable cell dehydrogenase
activity, the microculture tetrazolium cell proliferation assay
]. 20 μl of 5 mg/ml MTT (Sigma Aldrich, Poole, UK)
was added to each well for 3 h and the cells in each well
were solubilised in 150 μl of MTT solubilisation solution.
After overnight incubation, the optical density of each
well was measured on a 96-well plate reader (Dynatech,
Reading, UK) set at 570 nm wavelength. Known
concentrations of cells were also plated, cultured in the presence
of MTT, and similarly solubilized. The absorbance
reading of these control cells represents the metabolic activity
of a known number of cells and was used to generate a
standard curve in Microsoft Excel. The absorbance
reading for each sample (well) was directly compared with
the standard curve, and the numbers of viable cells were
Bystander killing effect studies
The bystander effect was determined by mixing HSV-TK
expressing cells with untransfected cells at the indicated
ratios. Cells were then plated in triplicate in 10 cm plates
at two densities, 1 × 105 and 5 × 105 cell/plate to ensure
cell–cell contact and to compare the in vitro effect of cell
densities on the bystander effect. Two days later, the cells
were treated with 50 μM GCV and incubated at 37 °C, 5%
CO2 for 10–14 days. The plates were then stained with
2% methylene blue and stained cells were counted.
In order to calculate the effect of GCV on the mixed
PA-STK and PA-1 populations, the number of colonies
counted were expressed as a percentage of the total
number of colonies in the co-cultured TK + ve and TK − ve
tumor cells at the indicated ratios of the two cell
populations in the presence of the indicated concentrations of
GCV. A graph was obtained by plotting the percentage of
γ‑irradiation of HSVT‑K modified tumor cells
2 expressed as 106 tumor cells resuspended in 5 ml of
DMEM were γ-irradiated (100 Gy) using a Gamma
cell1000 (Atomic Energy of Canada Ltd. Source: 137Cs).
Cytospin histological analysis
A 100 μl aliquot of 104 cells/ml in PBS was used to
prepare cytospin slides using a cytospin system (Hettich,
Salford, UK), fixed in 10% formalin for 10 min, left
overnight to air-dry and then stained by the Papanicolaou
staining method [
Measurement of apoptosis and necrosis by FACS analysis
Tumor cells were resuspended in 100 μl of working
labelling solution and incubated for 15 min in the dark. Two
control tubes were used: cells stained with Annexin
V-FITC (Becton–Dickinson, Oxford, UK) alone and cells
stained with PI alone [
]. The cells were then
analysed by flow cytometry using a FACScan analyser
(Becton–Dickinson, Oxford, UK).
Cell cycle analysis
Cells were irradiated at 100 Gy and incubated overnight
at 37 °C, in 5% CO2, resuspended in 1 ml of staining
buffer (0.1% sodium citrate, 0.1% Triton-X100 and 10 μg/
ml propidium iodide in dH2O), vortexed and left
overnight at 4 °C in the dark. The cells were then analysed by
flow cytometry using a FACScan analyser.
In vivo studies: inoculation, establishment and treatment of the tumors intraperitoneally (IP) in mice
AB1 mouse mesothelioma adherent tumor cells were
harvested by washing with versene only and re-suspended
in 0.2 ml of PBS. Tumors were established IP in female
BALB/c mice by injecting the mesothelioma cells (AB1
tumor cell line), at different doses (1 × 105, 5 × 105 and
1 × 106/100 ml PBS) using a 26-gauge needle, to
establish the TD50 for the tumor cell line, according to Home
Office regulations. Mice (n = 45) were injected i.p. with
AB1 tumor cells on day 0. Nine days later, animals were
assigned to nine groups (n = 5 per group). GCV
treatment was started at day 10. GCV (Cymevene® 500 mg;
Roche, Switzerland) was diluted in sterile DMEM to a
stock concentration of 50 mg/ml. The stock solution of
GCV was diluted in DMEM to a concentration of 2 mg/
ml, and 1 ml of the stock was injected IP once a day for
5 consecutive days. Mice were monitored every 2 days
to palpate the tumor. At post-mortem, all tumor
nodules were counted and measured using a calliper (for
each nodule, 2 perpendicular diameters were recorded).
Tumor volume was calculated for each nodule assuming
spherical shape and the total tumor volume was
calculated by adding all the calculated values for each mouse.
Statistical analysis was performed using the
Microsoft Excel program. Differences between groups were
analysed using Student’s paired t test. A P value of < 0.05
was considered as significant.
PA‑STK cells irradiated at 100 Gy lose their ability to induce bystander killing
Irradiation of PA-STK cells (100 Gy) substantially
reduced their ability to induce bystander killing of
unmodified PA-1 cells (Fig. 1a). This study was carried
out at the optimum number of 5 × 105 cells per 10 cm3
plate as previously determined (data not shown). A
possibility was that the irradiation stopped the growth of
PASTK cells and therefore reduced the possibility of cell to
cell contact in the tissue culture plate. This experiment
was therefore repeated at a higher cell density of 2 × 106
cells/plate. Increasing the cell density did not restore the
Ratios of PA-STK: PA-1 cells.
Ratios of PA-STK: PA-1 cells.
Ratios of AE17-STK: AE17 cells
Ratios of CRL5830-TK: CRL5830 cells
Fig. 2 In vitro bystander killing induced by γ-irradiated (100 Gy) mouse mesothelioma AE17-STK cells. a AE17-STK and AE-17 cells (with or without
γ-irradiation) were mixed and cultured in the presence of 50 μM GCV for 6 days. The total number of cells was 5 × 104/well of 96-well tissue culture
plate. Per cent survival was measured using the MTT assay. Each point is the mean of three separate measurements and error bars indicating
the standard error of the mean are shown. Similar data was obtained in three separate experiments. b In vitro bystander killing induced by the
γ-irradiated human mesothelioma cell line CRL-5830TK. CRL5830-STK and CRL5830 cells were mixed in the indicated ratios and cultured in the
presence of 50 μM GCV for 6 days. The total number of cells was 5 × 104/well of 96-well tissue culture plate. The fraction of surviving cells was measured
using the MTT assay. Each point is the mean of three separate measurements and error bars indicate the standard error of the mean. Similar data
was obtained in three separate experiments
bystander effect (Fig. 1b). At both cell densities,
irradiated PA-STK cells were highly significantly less efficient
at mediating the bystander effect at a 50:50 ratio than
unirradiated cells (P = 0.04). Similar data were obtained
in three further repeats of this experiment.
γ‑Irradiated (100 Gy) human and mouse mesothelioma
cells retain the ability to induce bystander killing
In contrast to the PA-STK cells, the mouse mesothelioma
AE17-STK cells retain their capacity to induce bystander
killing after γ-irradiation (100 Gy, Fig. 2a). Human
mesothelioma cells CRL-5830-TK, were similarly able to
retain their bystander killing activity after γ-irradiation
(Fig. 2b). In neither case was there a significant
difference in efficacy between irradiated and unirradiated cells
(P > 0.45 at all cell ratios).
Investigation of PA‑STK cell death after irradiation
Microscopic examination of the irradiated (100 Gy)
PASTK and PA-1 cells revealed that they were very sensitive
to irradiation. The cells did not attach to the culture plate
after irradiation, when visualised the next day (Fig. 3). In
contrast, OVC-432 cells were able to efficiently adhere to
the culture dish, even after 100 Gy γ-irradiation. These
cells, in common with all other cell lines tested
(OVC432, SKOV-3, OIB, CRL-5830, CRL-5839-STK,
CRL5820, CRL-5915, AE17, AE-STK, AB-1, AC-29, HL-60)
were able to attach efficiently to substrate and remain
metabolically active for about 3 days before eventually
losing surface adherence (data not shown).
A dose range of irradiation from 1 to 100 Gy was
then tested on PA-STK cells. For this purpose the cells
were trypsinised, γ-irradiated and then plated onto
tissue culture plates. The next day it was observed
that even the PA-STK cells irradiated at 1 Gy had lost
their ability to adhere to the tissue culture plates.
Further cell viability studies were performed using the
MTT assay to determine their viability, showing that
these cells were non-viable at all γ-irradiation doses.
Moreover, these cells failed to mediate any in vitro
bystander effect at all γ-irradiation doses tested (data
Analysis of cell death in PA‑STK cells
The first objective was to quantitatively record cell death
during a time course study following γ-irradiation using
the trypan blue dye exclusion method (Fig. 4). The
control cell line used in this experiment was another ovarian
cancer cell line, OVC-432, as well as non-irradiated
PASTK cells. The vast majority (over 90%) of the irradiated
PA-STK cells died by 18 h post irradiation. However, this
was not the case for control cells which survived beyond
Additionally, Annexin-V and propidium iodide staining
of cells was used as a method to quantify the proportion
of apoptotic and necrotic cells in the γ-irradiated
population of PA-STK cells, respectively. In this experiment
HL-60 cells treated with staurosporine (1 mg/ml for 4 h)
were used as positive control cells [
]. As shown in
Fig. 5, only a small proportion of PA-STK cells were dying
In order to further examine the mechanism of cell
death in the irradiated PA-STK cells, cytospin
preparations were examined 18 h later. The non-irradiated
PASTK cells served as controls. The cytospin histological
analysis of the irradiated PA-STK cells suggests that these
cells die by pyknosis, a form of necrosis (Fig. 6).
Pyknosis is characterised by nuclear shrinkage and increased
]. In the irradiated PA-STK cells the DNA
condensed into a solid, shrunken basophilic mass. The
only clear inference which can be drawn from these data
is that the radiation induced death in the vast majority
of PA-STK cells is due to necrosis/pyknosis, rather than
In vivo bystander killing activity
To establish an allogeneic TK-tumor cell/GCV
treatment model, a series of experiments were conducted to
repeat and extend previous in vivo studies. The
treatment groups in Fig. 6 were used to assess and compare
the activity of PA-STK cells (irradiated and
non-irradiated) as a possible vehicle for HSV-TK suicide gene
therapy with other TK expressing tumor cells. In this
experiment BALB/c mice were injected with the
syngeneic murine AB1 mesothelioma cells intraperitoneally
(1 × 106 cells per mouse) on day 0 (to act as the
experimental groups), and one group of mice acted as healthy
controls. Nine days later, the mice were separated into 9
different treatment groups. Group 1: PBS alone
intraperitoneally (2 ml), Group 2: AE17-STK alone (2 ml
containing 1 × 106 cells), Group 3: GCV alone (2 mg/ml/mouse
for 5 consecutive days), Group 4: GCV + AE17-STK
(GCV 2 mg/mouse for five consecutive days), Group 5:
γ-irradiated AE17-STK/GCV (100 Gy) + GCV, Group 6:
unirradiated PA-STK cells + GCV, Group 7: γ-Irradiated
PA-STK (100 Gy) + GCV, Group 8: CRL5830-STK/GCV,
and Group 9: γ-Irradiated CRL5830-STK/GCV (100 Gy).
Co-cultured cells for treatment were used at 50% HSVTK
modified tumor cells: 50% unmodified tumor cells. The
mice were then left and monitored for signs of tumor
development and toxicity. Eight days after the first day
of treatment (i.e. 17 days from tumor inoculation), mice
were sacrificed and post-mortem was performed. The
mean tumor volume was calculated for each treatment
group (Fig. 7).
Despite the toxicity of the treatment, the animals in
Group 4 (which received AE17-STK + GCV) had
significantly smaller tumors compared to the mice in the
first 3 control groups (PBS alone, AE17-STK alone,
or GCV alone) [P = 0.03]. This reduction in tumor
volume appeared to be reduced slightly in the case
of irradiated AE17-STK cells, as there was a smaller
reduction in tumor volume in Group 5 (which received
γ-irradiated AE17-STK with GCV) compared to Group
4 (GCV + AE17-STK), although this reduction was not
statistically significant (P = 0.56). For Groups 6 and
7 (PA-STK treated groups), the unirradiated PA-STK
cells + GCV mediated a detectable anti-tumor effect
in vivo, whereas the γ-irradiated PA-STK cells and GCV
did not (P values = 0.50 and 0.6 respectively), compared
to control mice which received PBS only. Moreover, both
the live and irradiated CRL5830-STK human
mesothelioma cells in combination with GCV mediated a
detectable anti-tumor bystander killing effect, compared to
control mice which received PBS only, P values = 0.05
and 0.06 respectively.
The use of lethally-irradiated allogeneic TK-modified
tumor cells could result in the development of a ‘generic’
HSV-TK expressing tumor cell line to generate an
antitumor immune response against a number of different
tumor types [
]. If effective, such a strategy could be
much more practical, as it could be more easily
standardised. In addition it could be a less expensive therapeutic
option than therapy with autologous modified tumor
The studies described above appear to indicate that
irradiation of PA-STK cells abolishes their bystander
killing ability, because the PA-STK cells are very
radiosensitive and die too rapidly to enable therapeutic benefit. The
radio-sensitivity of the PA-1 and PA-STK cells appeared
to be much higher than a panel of 10 other cell lines
including human and mouse mesothelioma cells. The
precise mechanism for this difference is unclear but could be
a result of a deficiency in DNA repair in PA-STK cells.
In a murine mesothelioma model, Schwarzenberger
and his colleagues successfully used PA-STK ovarian
tumor cells to treat peritoneally grown mesothelioma
tumor AC29. In this study the PA-STK cells were not
]. Efficacy of the irradiated TK + ve cells
was tested in a previous study by Freeman and his
colleagues using the TK + ve cells KBALB but not
PASTK cells. However, the data presented in this study
demonstrate that irradiation of PA-STK cells, even
at very low doses, makes them lose their capacity to
induce an in vitro bystander effect as they die very early
In this study, irradiated and non-irradiated AE17-STK
mouse mesothelioma cells were examined for their
efficiency in inducing the rejection of allogeneic tumors;
AE17-STK cells were also compared with a human
mesothelioma cell line CRL5830. Tumor regression was
detected in all animals injected with the non-irradiated
allogeneic AE17-STK cells after GCV administration
whereas in mice treated with the irradiated cells tumor
regression was detectable in only 60% of the animals.
Interestingly, the human mesothelioma CRL5830-STK
in combination with GCV treatment has indicated
significant reduction in tumor mass in contrast to PA-STK
human cell line.
The first clinical trial to use a suicide gene (HSV-TK)
as a primary therapeutic agent was approved in 1991 for
the treatment of ovarian carcinoma [
cancer cells (PA-1) were transfected ex vivo with the gene
for HSV-TK and then injected intraperitoneally in nine
patients with stage III disease. One patient achieved
complete remission, whereas others showed partial
tumor regression. However, data obtained in the present
study show that these particular cells (PA-STK) were not
able to mediate any detectable in vivo bystander
killing after lethal irradiation; however a detectable but not
significant bystander killing was found when these cells
were not irradiated. The high sensitivity of PA-STK to
irradiation appeared to be responsible for the complete
loss of the bystander effect not only in vivo but also
in vitro. The non-irradiated PA-STK cells were able to
mediate bystander killing in vitro, and also to induce a
detectable level of in vivo bystander killing. One possible
explanation is that the PA-STK cells are not very efficient
mediators of the direct bystander effect (PA-STK IC50 of
20 mM GCV, compared to an IC50 of 3 mM for
AE17STK or 1 mM for CRL5830-TK cells). It is possible that
the xenogeneic human ovarian PA-STK cells did not very
efficiently home onto and/or establish gap junctions with
the allogeneic murine mesothelioma cells [
However, as already mentioned, Schwarzenberger and
colleagues did find that PA-STK cells can treat AC29 mouse
mesothelioma tumors initiated 4 days earlier .
However, in the present studies in which the PA-STK cells
were given on day 9 of tumor inoculation there was no
evidence of tumor regression after irradiation, although
there was clear evidence for the induction of tumor
elimination by the AE17-STK cells.
To sum up, the issue of tumor cell sensitivity to
γ-irradiation is critical in cellular therapy, as ‘optimum’
sensitivity need to be observed carefully. Gene-modified
(HSVTK) tumor cells which are to be used in cancer
patients need to be lethally irradiated so that they will
not proliferate in the recipient’s body, but still be able
to mediate bystander killing. It is therefore important to
establish whether and when the irradiated gene-modified
tumor cells would die after irradiation.
This study suggests that PA-STK ovarian tumor cells
are rather poor mediators of bystander killing, and
therefore may not be the most efficient cells for future
clinical trials. However, the in vitro and in vivo results
obtained in the present studies suggest that CRL-5830TK
human mesothelioma cells may be effective
inducers of bystander killing for the clinical treatment of
mesothelioma. Interestingly, CRL-5830TK cells appear to
retain their killing ability even after lethal irradiation.
Although encouraging results are emerging from
preclinical and some early stage clinical studies, further
developments are needed. These include: (1)
development of relevant disease models in higher vertebrates
with physiological traits that are adequately
representative of humans malignancies; (2) development and
optimisation of the gene delivery systems; (3) further
investigation of ways of enhancing the bystander effect;
(4) modulation of the host immune response to enhance
and complement suicide gene therapy; (5) further
exploration of other therapeutic strategies in combination
with suicide gene therapy in the light of the observation
that certain suicide genes can behave as radio-sensitising
GCV: ganciclovir; HSV-TK: herpes simplex virus thymidine kinase; MTT:
microculture tetrazolium cell proliferation assay.
Jehad Zweiri conceived, planned and carried out the experiments and data
analyses, and drafted the manuscript; Steve Christmas advised on drafting the
manuscript. Both authors read and approved the final manuscript.
1 Department of Molecular and Haematological Medicine, King’s College
London, The Rayne Institute, 123 Coldharbour Lane, London SE5 9NU, UK.
2 Institute of Infection & Global Health, University of Liverpool, Liverpool, UK.
3 Dept. of Clinical Infection & Immunology, Institute of Infection & Global
Health, Ronald Ross Building, University of Liverpool, 8, West Derby St,
Liverpool L69 7BE, UK.
This study was conducted with the academic and financial support of
Professor Farzin Farzaneh of King’s College London, UK. Professor Farzaneh also
helped in the design of the experiments and reading the manuscript.
The authors declare that they have no competing interests.
Availability of data and materials
The data and materials of this study are included in this published article.
Ethics approval and consent to participate and Consent for publication
This manuscript contains no information on clinical materials and hence
ethical approval and consent for publication are not required.
This work was supported in part by King’s College London.
Springer Nature remains neutral with regard to jurisdictional claims in
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