[CANCER RESEARCH 52. 1764-1769. April 1, 1992|
Retention of Cellular Radiation Sensitivity in Cell and Xenograft Lines Established
from Human Melanoma Surgical Specimens'
Einar K. Rofstad2
Institute for Cancer Research and the Norwegian Cancer Society, Norwegian Radium Hospital, Montebello, 0310 Oslo 3, Norway
patients'
ABSTRACT
xenograft lines, e.g., karyotype, cell ultrastructure, and tumor
histology (17, 18). Experiments elucidating the validity of these
cell and xenograft lines as clinically relevant models in studies
of the radiation biology of malignant melanoma are reported
in this paper. The main purpose of the work was to investigate
to what extent the cellular radiation sensitivity of the donor
patients' tumors was retained during the heterotransplantation
Six human melanoma xenograft lines have been established in athymic
mice from metastatic lesions in six different patients. Permanent cell
lines in monolayer culture have been established from four of the xeno
graft lines. The cellular radiation sensitivity of the donor patients' tumors,
the xenograft lines, and the cell lines were measured in vitro. The
Courtenay soft agar colony assay was used for the donor patients' tumors,
and a conventional plastic surface colony assay was used for the cell
lines, whereas both assays were used for the xenograft lines. The cell
survival data were analyzed using the multitarget single-hit as well as
the linear-quadratic model. The donor patients' tumors differed consid
and the subsequent establishment of monolayer cell cultures.
Thus, survival curves were established in vitro for the cell and
xenograft lines and compared with those for cells isolated
directly from the donor patients' surgical specimens.
erably in cellular radiation sensitivity (the M, ranged from 0.85 ±0.08
to 1.17 ±0.09 Gy, the a from 0.25 ±0.06 to 0.87 ±0.14 Gy~', and the
surviving fraction at 2.0 Gy from 0.15 ±0.04 to 0.50 ±0.06). The
xenograft lines showed similar survival curves in soft agar and on the
plastic surface, and the survival curves of a xenograft line and the
corresponding cell line were not significantly different. These survival
curves were not significantly different from those of the donor patients'
tumors, regardless of which survival curve parameter was considered,
i.e., the cellular radiation sensitivity of the donor patients' tumors was
retained in the cell and xenograft lines. Moreover, the cell and xenograft
lines have growth properties in vitro and in vivo that render a wide
variety of experiments possible. Consequently, they show great promise
for future studies of human tumor radiation biology.
INTRODUCTION
Human tumor cell lines established in monolayer culture and
human tumor xenograft lines transplanted serially in athymic
mice are currently used as model systems in studies of the
radiation biology of human cancer (1-5). Such studies are often
based on the assumption that the cellular radiation sensitivity
of the donor patients' tumors is retained in the experimental
models (6-8). There is no unambiguous experimental evidence
that this assumption is valid (9). In fact, some studies have
indicated that the treatment sensitivity of tumor cells is changed
after establishment of permanent cell and xenograft lines (4, 5,
10, 11). Stress-induced genetic alterations as well as selection
of cell subpopulations during the establishment procedure are
possible causes of such changes (12, 13). Moreover, the radia
tion sensitivity of tumor cells may depend on the pretreatment
growth conditions and the assay used for measurement of
clonogenic cell survival (14-16).
Six human melanoma xenograft lines have recently been
established and characterized in our laboratory ( 17). Permanent
cell lines have been successfully established in monolayer cul
ture from four of the xenograft lines (18). The cell as well as
the xenograft lines were found to differ considerably with
respect to growth characteristics, physiological parameters, and
metastatic potential. Several biological features of the donor
Received 10/7/91 ; accepted 1/15/92.
The costs of publication of this article were defrayed in part by the payment
of page charges. This article must therefore be hereby marked advertisement in
accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
1Financial support was received from the Norwegian Cancer Society.
2 To whom requests for reprints should be addressed, at Department of
Biophysics. Institute for Cancer Research. Norwegian Radium Hospital. Monte
bello. 0310 Oslo 3, Norway.
tumors were shown to be retained in the cell and
MATERIALS AND METHODS
Mice. Eight- to 10-wk-old male BALB/c-nu/nu mice were used. They
were bred at the animal department of our institute and were kept
under specific-pathogen-free conditions at constant temperature (2426°C)and humidity (30 to 50%). Sterilized food and tap water were
given ml libitum.
Melanoma Lines. Six melanoma xenograft lines (BEX-t, COX-t,
HUX-t, ROX-t, SAX-t, WlX-t) have been established in athymic mice
from métastasesof patients (BEP-t, COP-t, HUP-t, ROP-t, SAP-t,
WIP-t, respectively) admitted to the Norwegian Radium Hospital (17).
Permanent cell lines (BEX-c, COX-c, SAX-c, WIX-c) have been estab
lished in monolayer culture from four of these xenograft lines (BEX-t,
COX-t, SAX-t, WlX-t, respectively) (18). The source tumors in the
donor patients, the xenograft lines, and the cell lines have been de
scribed in detail (17, 18). The donor patients did not receive radiation
therapy or preoperative chemotherapy (17).
The xenograft lines were maintained in athymic mice by serial s.c.
transplantation of tumor fragments, approximately 2x2x2
mm in
size (17). Subcutaneous flank tumors in passages 15 to 25, approxi
mately 500 mm1 in volume, were used in the present work. The lines
were kinetically stable during the period the experiments were carried
out, as ascertained by flow cytometric measurements of DNA histo
grams and measurements of volumetric growth rates.
The cell lines were maintained in 80-cm2 tissue culture flasks (Nunclon) to which were added 15 ml of RPMI 1640 medium (25 HIM
HEPES' and L-glutamine) which was supplemented with 13% fetal calf
serum, 250 mg/liter of penicillin, and 50 mg/liter of streptomycin (18).
They were incubated at 37°Cin a humidified atmosphere of 5% CO2
in air and subcultured once a week after trypsinization (treatment with
a 0.05% trypsin/0.02% EDTA solution for 2 min). Cells in passages
75 to 100 in vitro were used in the present work. Flow cytometric
measurements of DNA histograms and measurements of the rate of
cell proliferation showed that the lines were kinetically stable in these
passages. The cell lines were verified to be free from Mycoplasma
contamination by using both the Hoechst fluorescence and the mycoirim methods.
Single Cell Suspensions. Tissue specimens from the donor patients'
tumors were dropped into culture medium at 4°Cimmediately after
surgery and brought to the laboratory. Normal tissue, necrotic areas,
and blood clots were removed using a scalpel and a pair of tweezers.
The remaining tumor tissue was rinsed several times in cold culture
medium (4°C)and cut into fragments approximately 2x2x2
mm in
size. Some of the fragments were used to establish the xenograft lines
JThe abbreviations used are: HEPES. 4-(2-hydroxyethyl)-l-piperazineethanesulfonic acid; PE. plating efficiency; SF2, surviving fraction at 2.0 Gy.
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RADIATION SENSITIVITY OF TUMOR MODELS
(17), whereas single cell suspensions were prepared from others. Tissue
fragments were prepared from xenografted tumors using a similar
procedure.
A standardized mechanical procedure was used to prepare single cell
suspensions from the fragments, whether derived from the donor pa
tients' tumors or from xenografted tumors. The fragments were put
into a plastic bag with 20 ml of culture medium and disaggregated for
30 s with a stomacher (Lab-Blender 80). The resulting suspension was
filtered through 30-Mm nylon mesh to remove cell aggregates, centrifuged, and resuspended in complete culture medium [Ham's F-12
medium supplemented with 20% fetal calf serum, 250 mg/liter of
penicillin, and 50 mg/liter of streptomycin for growth in soft agar or
RPMI 1640 medium (25 mM HEPES and L-glutamine) supplemented
with 13% fetal calf serum, 250 mg/liter of penicillin, and 50 mg/liter
of streptomycin for growth in monolayer culture]. The quality of the
suspensions was examined using a phase-contrast microscope and a
hemocytometer. The fraction of doublets was always <5%, and the
fraction of larger aggregates was always <0.1%. The number of mor
phologically intact, metabolically viable cells, i.e., cells having an intact
and smooth outline with a bright halo, were counted, and the cell
suspensions were diluted to appropriate concentrations for use in
radiation experiments.
Irradiation. A Siemens Stabilipan X-ray unit, operated at 220 kV, 20
mA, and with 0.5-mm Cu filtration, was used for irradiation. The
irradiation was performed at room temperature at a dose rate of 3.4
Gy/min. The cells were kept in 1 ml of soft agar or liquid culture
medium in plastic tubes (Falcon 2057) during the radiation exposure.
Cell suspensions from tumors were prepared as described above. Cell
suspensions from monolayer cultures were obtained by trypsinization
of cultures in exponential growth. Exponential growth was achieved by
plating 5 x IO4cells in 80-cm2 tissue culture flasks and harvesting the
cells after 4 to 6 days (18).
Soft Agar Clonogenic Assay. The soft agar colony assay used in the
present work was developed by Courtenay and Mills (19). The soft agar
was prepared from powdered agar (Bacto agar; Difco) and culture
medium (Ham's F-12 medium supplemented with 20% fetal calf serum,
250 mg/liter of penicillin, and 50 mg/liter of streptomycin). Erythrocytes from August rats were added before melanoma cells were em
bedded in the soft agar. Aliquots of 1 ml of the soft agar were seeded
in plastic tubes (Falcon 2057). The tubes were plunged into crushed
ice, and the soft agar was allowed to set.
The cells were incubated at 37°Cfor 4 to 5 wk in an atmosphere of
5% O2, 5% CO2, and 90% N2. Culture medium (2 ml) was added to the
top of the soft agar 5 days after seeding and then changed weekly.
Colonies were counted using a stereomicroscope. Cells giving rise to
colonies containing more than 50 cells were scored as clonogenic. The
PE was calculated from the number of colonies counted and the number
of morphologically intact, metabolically viable cells seeded.
»o
Previous work has shown that there are potential pitfalls in this soft
agar colony assay when used to study survival of cells isolated directly
from human tumors (20, 21). Care was taken to avoid these pitfalls.
(a) The relationship between number of colonies and number of cells
seeded was verified to be linear for all donor patients' tumors and
xenograft lines, (b) Reliable criteria to distinguish vital, real colonies
from large abortive colonies were established. These criteria were based
on examinations of the size, shape, and color of the colonies from
unirradiated cells (20). (c) Scoring of artificial colonies due to cell
clumps was avoided by examining soft agar cultures of cells given a
lethal radiation dose of 30.0 Gy. (d) Finally, measured surviving frac
tions were corrected mathematically for multiplicity before curve fitting
was performed (22).
Plastic Surface Clonogenic Assay. Melanoma cells, suspended in 1
ml of RPMI 1640 culture medium (25 m.MHEPES and L-glutamine)
supplemented with 13% fetal calf serum, 250 mg/liter of penicillin, and
50 mg/liter of streptomycin, were plated in 25-cm2 tissue culture flasks
(Nunclon). The flasks contained 1 x IO5 lethally irradiated (30.0 Gy)
feeder cells in 4 ml of culture medium, which were plated 24 h earlier.
It was verified experimentally that the use of feeder cells increased the
PE, particularly for cells isolated directly from xenografted tumors.
Moreover, a linear relationship between number of colonies and number
of cells seeded was ensured by the use of feeder cells. Finally, the use
of feeder cells inhibited migration of viable cells, hence causing dense
and easily scorable colonies.
The cells were incubated at 37°Cfor 1 to 3 wk in a humidified
atmosphere of 5% CO: in air. One half of the culture medium (2.5 ml)
was removed and replaced with fresh medium each week. The cells
were fixed in 100% ethanol and stained with méthylène
blue. Colonies
containing more than 50 cells were counted using a stereomicroscope.
Surviving fractions were calculated and corrected for multiplicity in the
same way as in the soft agar colony assay.
Curve Fitting and Statistical Analysis. Survival curves were fitted to
the data by the method of least squares using the multitarget single-hit
as well as the linear-quadratic model. The curve fitting was based on
surviving fractions from individual experiments as opposed to mean
values. A two-tailed / test was used to investigate whether the parame
ters of two survival curves were significantly different.
RESULTS
Radiation survival curves in vitro for the donor patients'
tumors and the corresponding cell and xenograft lines are
presented in Figs. 1 to 6. The donor patients' tumors were
assayed for cell survival in soft agar, whereas the plastic surface
assay was used for the cell lines. The four xenograft lines from
which cell lines were successfully established were assayed both
10"
100
BEP-I
soft ag
10"
BEX-1
soft ogci
BEX-c
pest*
10-1
10-3
10'
6
6
X) 12
02
X) 12 0 2
8
10
12
8
10 12
2
DOSE (Gy)
Fig. 1. Radiation survival cunes for BEP-t, BEX-t. and BEX-c human melanoma cells irradiated in vitro. The BEP-t cells were assayed for survival in soft agar.
the BEX-c cells on plastic surface in conventional tissue culture flasks, and the BEX-t cells both in soft agar and on plastic surface. Points, surviving fractions
calculated from the mean number of colonies in 4 soft agar tubes or 4 tissue culture flasks. Different points at the same dose level refer to independent experiments.
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RADIATION SENSITIVITY OF TUMOR MODELS
D"Z
10°
COP-I
san
agoI\i
\.
agotxL
COX-1
pusu
soft
V\.
IO'1*
- 10-'
\vÀ10Ux)-'Xi'2X)-3«t\COX-t
\\:\v
XT'115?
- 10"
-2
10
-J
«-3\
-
10"
10-3
X)'
0
2
4
6
8
K>
12
0246810
12
0246810
12
10
02468
10
12
DOSE (Gy)
Fig. 2. Radiation survival curves for COP-t, COX-t, and COX-c human melanoma cells irradiated in vitro. The COP-t cells were assayed for survival in soft agar,
the COX-c cells on plastic surface in conventional tissue culture flasks, and the COX-t cells both in soft agar and on plastic surface. Points, surviving fractions
calculated from the mean number of colonies in 4 soft agar tubes or 4 tissue culture flasks. Different points at the same dose level refer to independent experiments.
in soft agar and on plastic surface, whereas only the soft agar
assay was used for the other two xenograft lines. The cell
survival curve parameters, SF2, and PE are presented in
Table 1.
The BEX-t and COX-t lines showed a higher PE on plastic
surface than in soft agar, whereas the PE was similar in these
two assays for the SAX-t and WlX-t lines. The HUX-t and
ROX-t lines were not able to form colonies on plastic surface.
The PE of the xenograft lines was somewhat higher or similar
to that of the corresponding donor patients' tumors. The cell
lines always showed a higher PE than did the xenograft lines.
The xenograft lines showed similar survival curves in soft agar
and on plastic surface, i.e., the cell surviving fractions did not
vary with the in vitro assay. The survival curves of a xenograft
line and the corresponding cell line were not significantly
different, even though the PE (Table 1) and the fraction of cells
in S phase (17, 18) were lower in the xenograft lines than in
the cell lines, suggesting that the survival curves were insensitive
to variations in these parameters. Moreover, none of these
survival curves differed significantly from those of the donor
patients' tumors. This conclusion was found to be valid, no
matter which of the survival curve parameters in Table 1 were
considered in the statistical analysis. Consequently, the cellular
radiation sensitivity of the donor patients' tumors was retained
in the cell and xenograft lines.
The cellular radiation sensitivity differed considerably among
the different donor patients' tumors, xenograft lines, and cell
lines. This is illustrated in Fig. 7, where the soft agar survival
curves for the xenograft lines are redrawn in the same panel.
The ROX-t was the most resistant and the BEX-t the most
sensitive line. The a and SF2, for example, varied within a
factor of about 3 among the lines. The xenograft lines have also
been found to differ significantly in several growth and phys
iological parameters in vivo (17). There was no relationship
between the cellular radiation sensitivity and these parameters.
This observation, together with the observation that the donor
patients' tumors and the corresponding cell and xenograft lines
showed similar survival curves, suggests that the differences in
cellular radiation sensitivity in Fig. 7 were not merely a second-
100
O
ÃŽ
i
o
-3
«r*
KT*
«T*
8
X)
12
8
10
8
12
DOSE (Gy)
Fig. 3. Radiation survival curves for HUP-t and HUX-t human melanoma
cells irradiated in vitro and assayed for survival in soft agar. Points, surviving
fractions calculated from the mean number of colonies in 4 soft agar tubes.
Different points at the same dose level refer to independent experiments.
10
12
8
K)
DOSE (Gy)
Fig. 4. Radiation survival curves for ROP-t and ROX-t human melanoma cells
irradiated in vitro and assayed for survival in soft agar. Points, surviving fractions
calculated from the mean number of colonies in 4 soft agar tubes. Different points
at the same dose level refer to independent experiments.
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RADIATION SENSITIVITY OF TUMOR MODELS
10»
TO"
10°
Kft
SAP-1
soft ogor
10•¿
«T»
10'
8
C
12
0246
8
t>
12
02
DOSE
8
10
6
12
8
10
12
(Gy)
Fig. 5. Radiation survival curves for SAP-t, SAX-t, and SAX-c human melanoma cells irradiated in vitro. The SAP-t cells were assayed for survival in soft agar,
the SAX-c cells on plastic surface in conventional tissue culture flasks, and the SAX-t cells both in soft agar and on plastic surface. Points, surviving fractions
calculated from the mean number of colonies in 4 soft agar tubes or 4 tissue culture flasks. Different points at the same dose level refer to independent experiments.
10°
v
I
WIX-I
otaste
WIX-I
soft agar
WIP-I
soft ogor
XT'
Õ
XT'
IO-'
0-3
10-3
Kf
81012
8t)
12
02
8
10
12
468
10
12
DOSE (Gy)
Fig. 6. Radiation survival curves for WIP-t, WlX-t, and WIX-c human melanoma cells irradiated in vitro. The WIP-t cells were assayed for survival in soft agar,
the WIX-c cells on plastic surface in conventional tissue culture flasks, and the WlX-t cells both in soft agar and on plastic surface. Points, surviving fractions
calculated from the mean number of colonies in 4 soft agar tubes or 4 tissue culture flasks. Different points at the same dose level refer to independent experiments.
ary effect, but were rather a direct consequence of genetic
different for tumors in humans, xenografted tumors, and cells
in culture (4, 30).
The six human melanoma xenograft lines studied here were
all found to have retained the cellular radiation sensitivity of
the donor patients' tumors. Moreover, the four cell lines that
;rences among the lines.
DISCUSSION
Experiments performed with human tumor xenograft lines
suffer from the uncertainty of whether the xenografted tumors
may have other biological properties and treatment responses
than the source tumors in the donor patients (4, 5, 23, 24). The
same uncertainty also applies to continuous cell lines estab
lished in monolayer culture (10, 11, 25). Changes in radiation
sensitivity may occur through at least three fundamentally
different mechanisms when tumor cells are removed from the
host and cultured in an artificial environment /// vivo or in vitro,
(a) Many tumors consist of cell subpopulations that can differ
in several biological properties, including radiation sensitivity
(26, 27). Tumor cells are usually subjected to a strong selection
pressure in a new environment, which may lead to complete
dominance of one of the subpopulations (12). (b) Tumor cells
are often genetically unstable, and extensive genotypic and
phenotypic changes may occur during growth in a new environ
ment (13, 28). (c) Finally, the radiation sensitivity of tumor
cells depends on the cell cycle distribution and the physiological
conditions in the microenvironment (14, 29), which often are
were successfully established in monolayer culture showed sim
ilar radiation sensitivity to the source xenograft lines. Conse
quently, processes leading to changes in cellular radiation sen
sitivity, such as the processes described above, do probably not
occur very frequently when cell and xenograft lines are estab
lished from human melanoma métastases.However, there is
clear evidence from studies of other melanomas that changes
in radiation sensitivity indeed can take place during establish
ment of cell lines in vitro. Studies in our laboratory showed that
the cell line FME established from the human melanoma xen
ograft line E.F. was more sensitive to radiation treatment than
the source xenograft line. FME cells in exponential growth
showed a D0 of 0.68 ±0.03 Gy, whereas the Da of the E.F. cells
irradiated in vitro was 1.32 ±0.08 Gy (10).
The individual donor patients' tumors, xenograft lines, and
cell lines studied here differed considerably in cellular radiation
sensitivity. This observation is in agreement with results pre
sented in a recent review of the radiation biology of malignant
melanoma (31). The review concluded that the differences in
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RADIATION SENSITIVITY OF TUMOR MODELS
Table 1 Radiation survival curve parameters
(Gy-2)0.041
(Gy-)0.87
CellsBEP-tBEX-tBEX-tBEX-cCOP-tCOX-tCOX-tCOX-cHUP-tHUX-tROP-tROX-tSAP-tSAX-tSAX-tSAX-cWIP-tWlX-tWIXtWIX-cAssaySoft
(Gy)0.85
0.08f0.93
±
agarSoft
±0.71.3
agarPlasticPlasticSoft
0.070.83
±
±0.31.8
0.060.83
±
±0.61.6
0.44.7
+
0.040.84
±
agarSoft
±0.120.89
5.83.6
±
agarPlasticPlasticSoft
0.050.88
±
1.73.9
±
0.040.84
±
1.54.7
±
2.56.5
±
0.060.92
±
0.060.99
±
3.24.3
±
agarSoft
agarSoft
±0.101.17
3.55.3
±
0.091.10
+
2.57.7
±
agarSoft
3.17.8
+
±0.060.99
agarSoft
5.27.0
±
agarSoft
0.091.02
±
3.18.0
+
±0.071.00
agarPlasticPlasticSoft
±0.070.94
3.911.8
±
±4.03.4
0.040.97
±
5.03.7
±
agarSoft
0.220.95
±
agarPlasticPlasticDo
0.070.85
±
±2.19.8
3.72.9+
+
0.040.98
±
1.1«
±0.04n1.8
" Values calculated from the linear-quadratic curve fit.
* Values measured in vitro.
c Mean + SE.
0.140.87
±
0.130.79
+
0.130.94
+
±0.110.58
0.110.69
+
0.080.65
±
0.070.66
±
0.100.38
+
0.050.48
±
0.120.25
+
0.060.29
±
0.070.27
±
0.080.28
±
0.080.27
±
0.080.30
±
0.060.40
+
0.130.55
±
0.090.34
+
0.050.59
±
±0.06ß
(%)1-31-310-3090-1005-1010-30
0.0290.032
+
0.040.1
+
±0.030.14
0.020.14
+
0.0250.055
±
±0.040.17
5
0.0210.034
±
±0.040.13
±0.030.14
0.030.25
+
170.058±0.0
±0.030.24
0.0180.035
+
0.050.22
+
0.040.22
+
0.0090.041
+
0.030.23
+
0.040.23
±
0.0080.042
+
0.040.23
+
0.040.25
±
0.050.37
+
0.0120.061
+
0.040.39
+
0.0070.046
±
0.040.32
±
0.030.39
±
0.070.50
+
0.0140.050
±
0.030.51
±
0.060.47
±
0.0080.044
±
±0.050.52
0.0070.060
+
0.070.46
±
0.030.45
±
0.0100.057
+
0.070.45
±
0.030.44
±
0.0100.060
+
0.060.46
+
0.040.49
±
0.060.47
±
±0.0100.054
0.070.44
±
0.0050.073
±
0.050.34
±
0.040.40
±
0.080.28
+
0.040.35
+
0.0210.045
+
110.072±0.0
0.050.38
±
0.050.45
±
0.030.37
+
0.050.26
+
0.0070.040
±
+ 0.04SFj*0.17
+ 0.02PE
±0.007SF2"0.15
without the use of enzymes, and the cells can be plated and
grown in vitro. Cells from the established lines form tumors
which are histologically indistinguishable from those of the
source xenograft lines when reinjected into athymic mice (18).
The PE is sufficiently high for most purposes both in soft agar
and on plastic surface. The cell survival curves are insensitive
to variations in PE and cell cycle distribution. The cellular
radiation sensitivity is similar in the soft agar and the plastic
surface assay. Consequently, the six xenograft lines and the
four cell lines constitute an experimental tumor panel that
shows great promise for future studies of human tumor radia
tion biology in general and the radiation biology of malignant
melanoma in particular.
o
gu_
ACKNOWLEDGMENTS
O
2
¿
6
8
K)
The skillful technical assistance of Berit Mathiesen, Hanne Stageboe
Petersen, and Heidi Kongshaug is gratefully acknowledged.
12
DOSE (Gyl
Fig. 7. Radiation survival curves for BEX-t, COX-t. HUX-t, ROX-t, SAX-t,
and WlX-t human melanoma cells irradiated in vitro and assayed for survival in
soft agar. The curves are redrawn from Figs. 1 to 6 for comparison.
REFERENCES
1. Fértil,B., and Malaise, E. P. Intrinsic radiosensitivity of human cell lines is
radiation sensitivity among melanoma cell lines irradiated in
vitro can be almost as large as those among cell lines from
tumors of different histology and with very different clinical
radiocurability (31). Moreover, recent clinical investigations
also suggest that malignant melanoma no longer should be
considered to be a typically radiation-resistant tumor type (32,
33). The data rather indicate that melanomas are heterogeneous
in clinical radioresponsiveness; some tumors may be resistant
but others are sensitive.
Clinically relevant radiobiological studies require adequate
experimental tumor model systems. A panel of cell and xeno
graft lines established from different patients suffering from the
same histopathological type of neoplastic disease is described
here. These lines constitute a particularly useful model system
for radiobiological studies, because they differ considerably in
cellular radiation sensitivity and because the cellular radiation
sensitivity of the donor patients' tumors was retained during
the establishment procedure in vivo and in vitro. In addition,
the lines have growth properties that render a wide variety of
experiments possible. Xenografted tumors can be disaggregated
correlated with radioresponsiveness of human tumors: analysis of 101 pub
lished survival curves. Int. J. Radiât.Oncol. Biol. Phys., //: 1699-1707,
1985.
Deacon, J., Peckham, M. J., and Steel, G. G. The radioresponsiveness of
human tumours and the initial slope of the cell survival curve. Radiother.
Oncol., 2:317-323, 1984.
Elkind, M. M. The initial part of the survival curve: does it predict the
outcome of fractionated radiotherapy? Radiât.Res., 114:425-436, 1988.
Rofstad, E. K. Radiation biology of human tumour xenografts. Int. J. Radiât.
Biol., 56:573-581, 1989.
Sutherland, R. M., Rasey, J. S., and Hill, R. P. Tumor biology. Am. J. Clin.
Oncol., //: 253-274, 1988.
Peters, L. J., Brock, W. A., Chapman, J. D., and Wilson, G. Predictive assays
of tumor radiocurability. Am. J. Clin. Oncol., li: 275-287, 1988.
Steel, G. G., McMillan, T. J., and Peacock, J. H. The picture has changed
in the 1980s. Int. J. Radiât.Biol., 56: 525-537, 1989.
Weichselbaum. R. R., Rotmensch, J., Ahmed-Swan, S., and Beckett, M. A.
Radiobiological characterization of 53 human tumor cell lines. Int. J. Radiât.
Biol., 56: 553-560, 1989.
Rofstad, E. K., Zaffaroni, N., and Hystad, M. E. Heterogeneous radiation
and heat sensitivity in vitro of human melanoma xenograft lines established
from different lesions in the same patient. Comparisons with the radiation
and heat sensitivity of cells isolated from the donor patient's surgical speci
mens. Int. J. Radial. Biol., 57: 1113-1122, 1990.
10. Rofstad, E. K., and Brustad, T. Radiosensitivity of the cells of an established
human melanoma cell line and the parent melanoma xenograft. Int. J. Radiât.
Biol., 44:447-454. 1983.
Tveit, K. M., Fodstad, 0., and Pihl, A. The usefulness of human tumor cell
1768
Downloaded from cancerres.aacrjournals.org on July 31, 2017. © 1992 American Association for Cancer Research.
RADIATION SENSITIVITY OF TUMOR MODELS
12.
13.
14.
15.
16.
17.
18.
19.
20.
21.
lines in the study of chemosensitivity. A study of malignant melanomas. Int.
J. Cancer, 28: 403-408, 1981.
Heppner, G. H. Tumor heterogeneity. Cancer Res., 44: 2259-2265, 1984.
Hill, R. P. Tumor progression: potential role of unstable genomic changes.
Cancer Metastasis Rev., 9: 137-147, 1990.
Kwok, T. T., and Sutherland, R. M. The relationship between radiation
response of human squamous carcinoma cells and specific metabolic changes
induced by chronic hypoxia. Int. J. Radial. Oncol. Biol. Phys., 16: 13011305, 1989.
Rofstad, E. K. Growth and radiosensitivity of malignant melanoma multicellular spheroids initiated directly from surgical specimens of tumours in
man. Br. J. Cancer, 54: 569-578, 1986.
West, C. M. L., and Sutherland, R. M. A radiobiological comparison of
human tumor soft-agar clonogenic assays. Int. J. Cancer. 37:897-903, 1986.
Rofstad, E. K.. Wahl, A., Stokke. T.. and Nesland, J. M. Establishment and
characterization of six human melanoma xenograft lines. Acta Pathol. Mi
crobiol. Immunol. Scand., 98: 945-953, 1990.
Rofstad, E. K., Wahl, A., Hystad, M. E., Nesland, J. M.. and Stokke. T.
Establishment in monolayer culture and characterization of four human
melanoma cell lines. Virchows Arch. B Cell Pathol., 60: 189-197, 1991.
Courtenay, V. D., and Mills, J. An in vitro colony assay for human tumours
grown in immune-suppressed mice and treated in vivo with cytotoxic agents.
Br. J. Cancer, 37: 261-268, 1978.
Rofstad, E. K., Wahl, A., Tveit, K. M., Monge, O. R., and Brustad, T.
Survival curves after X-ray and heat treatments for melanoma cells derived
directly from surgical specimens of tumours in man. Radiother. Oncol., 4:
33-44, 1985.
Rofstad, E. K.. Wahl, A., and Brustad, T. Radiation sensitivity in vitro of
cells isolated from human tumor surgical specimens. Cancer Res., 47: 106110, 1987.
22. Gillespie, C. J., Chapman, J. D., Reuvers, A. P., and Dugle, D. L. The
inactivation of Chinese hamster cells by X rays: synchronized and exponential
cell populations. Radiât.Res., 64: 353-364, 1975.
23. Steel, G. G., Courtenay, V. D., and Peckham, M. J. The response to
chemotherapy of a variety of human tumour xenografts. Br. J. Cancer, 47:
1-13, 1983.
24. Giovanella. B. C., and Fogh, J. The nude mouse in cancer research. Adv.
Cancer Res., 44: 69-120, 1985.
25. Connors, T. A., and Phillips, B. J. Screening for anticancer agents. The
relative merits of in vitro and in vivo techniques. Biochem. Pharmacol., 24:
2217-2224,1975.
26. Leith, J. T., Dexter, D. L., DeWyngaert, J. K., Zeman, E. Z., Chu, M. Y.,
Calabresi, P., and Glicksman. A. S. Differential responses to X-irradiation
of subpopulations of two heterogeneous human carcinomas in vitro. Cancer
Res., 42:2556-2561, 1982.
27. Rofstad, E. K., and Brustad, T. Differential responses to radiation and
hyperthermia of cloned cell lines derived from a single human melanoma
xenograft. Int. J. Radial. Oncol. Biol. Phys., 10: 857-864. 1984.
28. Nicolson, G. L. Tumor progression, oncogenes, and the evolution of metastatic phenotypic diversity. Clin. Exp. Metastasis, 2: 85-105, 1984.
29. Denekamp, J. Cell kinetics and radiation biology. Int. J. Radiât.Biol., 49:
357-380, 1986.
30. Moulder, J. E.. Dutreix, J., Rockwell, S., and Siemann, D. W. Applicability
of animal tumor data to cancer therapy in humans. Int. J. Radial. Oncol.
Biol. Phys., 14: 913-927. 1988.
31. Rofstad. E. K. Radiation biology of malignant melanoma. Acta Radiol.
Oncol.. 25: 1-10. 1986.
32. Habermalz, H. J. Irradiation of malignant melanoma. Experience in the past
and present. Int. J. Radial. Oncol. Biol. Phys., 7: 131-133, 1981.
33. Harwood, A. R.. and Cummings. B. J. Radiotherapy for malignant mela
noma. A re-appraisal. Cancer Treat. Rev.. 8: 271-282, 1981.
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Retention of Cellular Radiation Sensitivity in Cell and Xenograft
Lines Established from Human Melanoma Surgical Specimens
Einar K. Rofstad
Cancer Res 1992;52:1764-1769.
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