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The performance status scale for head and neck cancer patients and the functional assessment of cancer therapy-head and neck scale A study of utility and validity

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2280
Analysis of Response to Radiation Therapy of Patients
with Cervical Adenocarcinoma Compared with
Squamous Cell Carcinoma
M I 6 4 and PClO Labeling Indices
Kuniyuki Oka, M.D.'"
Takashi Nakano, M.D?
Tanji Hoshi, M.D."
' Department of Pathology, Mito Saiseikai General Hospital, Mito, Ibaraki, Japan.
Unit of Clinical Examinations, National Institute of Radiological Sciences, Chiba, Japan.
Unit of Clinical Oncology, 2,3. Division of Radiation Medicine, Research Center of Charged
Particle Therapy, National Institute of Radiological Sciences, Chiba, Japan.
Institute for Urban Studies, Tokyo Metropolitan University, Tokyo, Japan.
This study was supported by a grant for the
diagnostic study group of charged particle therapy project (National Institute of Radiological
Sciences; NIRS, Chiba, Japan).
The authors thank Mr. 0. Mrozekfor the English
revision, Dr. A. Tsuboi for helpful suggestions
and advice, Mr. R. Ohuchi (NIRS), Mr. Y. Naoi,
Mrs. S. Koyamatsu, Mr. S. Nakamura, and Miss
T. Tobita (Mito Saiseikai General Hospital) for
excellent technical support, and Mrs. M. Sakurai
(NIRS) for secretarial assistance.
Address for reprints: Kuniyuki Oka, M.D.,Department of Pathology, Mito Saiseikai General
Hospital, 3-3-10 Futabadai, Mito, lbaraki 31 141, Japan.
Received December 18, 1995; accepted February 9, 1996.
0 1996 American Cancer Society
BACKGROUND. The MIB-1 monoclonal antibody is a marker of cycling cells and
the PClO monoclonal antibody is a marker of proliferating cell nuclear antigen in
paraffin sections. This study was conducted to elucidate the difference in response
to radiotherapy (RT) between cervical adenocarcinomas and squamous cell carcinomas, focusing on cell proliferation.
METHODS. A total of 196 biopsy specimens taken from the cervical carcinomas of
14 consecutive patients with adenocarcinoma and 62 patients with squamous cell
carcinoma before and after RT at doses of 9 and 27 Grays (Gy) were investigated
for MIB-1 and PClO immunoreactivities.
RESULTS. In adenocarcinomas, the mean MIB-1 labeling indices before and after
RT at 9 and 27 Gy were 28%, 21%, and 26%, respectively, whereas the mean
PClO labeling indices were 15%, 13%, and 14%, respectively. In squamous cell
carcinomas, the mean MIB-1 labeling indices before and after RT at 9 and 27 Gy
were 38%, 53%, and 26%, respectively, and the mean PClO labeling indices were
23%, 23%, and 11%, respectively.
CONCLUSIONS. Cervical adenocarcinomas have a lower cycling cell population and
their indices show no change during RT. Squamous cell carcinomas have a higher
cycling cell population and show a transient increase of the MIB-1 cycling cell population at 9 Gy of RT. These findings suggest a difference in response to RT between
adenocarcinomas and squamous cell carcinomas. Cancer 1996; 77:2280-5.
0 1996 American Cancer Society.
KEYWORDS: radiation, cell cycle, MIB-1 , PC10, cervical carcinoma, immunohistochemistry.
I
n general, the prognosis of adenocarcinomas of the cervix is regarded
as poorer than that of squamous cell carcinomas'z2;two reasons for this
are poorer local control rate and high incidence of metastasis. The poorer
local control rate is thought to be due to the lower radiosensitivity compared with squamous cell carcinomas. Although the nature and clinical
characteristics of squamous cell carcinomas have been studied extensively, the features of adenocarcinomas of the cervix are not yet fully
understood, due to their low incidence. The mechanism behind the low
radiation sensitivity of adenocarcinomas is a major concern and requires
further investigation.
Recently, many types of molecules associated with the cell cycle,
including proliferating cell nuclear antigen (PCNA), Ki-67 antigen, and
p53 oncoprotein, have enabled us to elucidate the proliferative status of
human
The growth fractions, proliferating cell nuclear antigen
Response to Radiation in Cervical CarcinornalOka et al.
(PCNA), and mitotic indices of squamous cell carcinomas
of the uterine cervix have been intensively investigated by
several groups of researchers, including our groups,"" and
a transient increase in growth fraction has been reported
in relation to the radiation response of the tumor cell^.^^'"
However, the cell cycle dynamics of adenocarcinomas of
the cervix during radiotherapy (RT) have not been evaluated. Analysis of the cell cycle in adenocarcinomas may
elucidate the mechanism responsible for its low radiation
sensitivity.
The purpose of the present study, therefore, was to
assess the tumor proliferation status of adenocarcinomas
of the uterine cervix before and during RT in comparison
with that of squamous cell carcinomas.
METHODS
We investigated I 4 patients with cervical adenocarcinornas who received a single established RT protocol at the
National Institute of Radiological Sciences Hospital
(Chiba, Japan). Their ages ranged from 36 to 82 years
(mean, 65 years). Five patients had Stage IIB disease, six
had Stage IIIb disease, and three had Stage IVA disease.
Six patients had adenocarcinoma of the endocervical
type, 2 had endometrioid adenocarcinoma, and six had
adenosquamous carcinoma. We also investigated 62 paiients with cervical squamous cell carcinomas who received RT. Their ages ranged from 35 to 83 years (median,
63 years). Three patients had Stage I disease, 17 had Stage
IIB disease, 38 had Stage IIIb disease, and 4 had Stage
1Va disease. Thirteen patients had squamous cell carcinoma of keratinizing cancer, 36 had large cell nonkeratinizing cancer, and 13 had small cell nonkeratinizing cancer.
Radiotherapy Protocol
The patients were treated with a combination of external
beam RT and high dose rate remote afterloading intracavitary RT." The external pelvis RT was administered as
10-megaelectron volt photon beams or cobalt-60 gamma
rays, with a dose of 1.8 Gray (Gy) per fraction, 5 times
per week. The intracavitary RT was delivered by a Ralstron system (Shimazu Corporation, Tokyo, Japan) which
has high dose rate cobalt-60 sources, with 2-4 Ci as the
intracavitary source. The details of the RT procedure have
been described previou~ly.~~"
Histopathologic Study
Prior to RT, biopsy specimens were obtained for the
pathologic diagnosis of cervical tumors of all 14 patients
with adenocarcinomas. Subsequently, one or two biopsy
specimens were obtained from the cervical tumors during
the period of RT, 1 day after total whole-pelvis irradiation
of 9 Gy (n = 9) and/or 27 Gy (n = 7). Specimens were
also obtained from the 62 patients with squamous cell
2281
carcinomas for the pathologic diagnosis of the cervical
tumors before RT. Subsequently, one or two biopsy specimens were obtained from the cervical tumors during the
period of RT 1, day after radiation at 9 Gy (n = 62) and1
or 27 Gy (n = 42). The intervals between the initiation of
radiation and the biopsy at radiation doses of 9 and 27
Gy were 8 and 22 days, respectively.
A total of 196 specimens were fixed with 10%formaldehyde for approximately 24 hours, and embedded in
paraffin for hematoxylin and eosin staining and MIB-1
and PClO immunostaining. MIB-1 antibody recognizes
Ki-67 antigen in paraffin section^.^,^ The MIB-1 labeling
index represents the cycling cell population, this being
equivalent to the growth fraction. PCNA is a nuclear protein associated with the cell cycle and DNA synthesis.
Anti-PC10 antibody recognizes PCNA in paraffin sections,
and the PClO labeling index mainly represents the Sphase fraction."
For MIB- 1 immunostaining, the sections were cut
and placed on glass slides coated with 3-aminopropyltriethoxysilane (LS-3150; Silicon Chemicals, ShinEtsu, Tokyo,
Japan), after which they were heated 8 times in a household microwave oven (Mitsubishi Electric System Ovenrange RO-3700, Tokyo, Japan), each time for 3 minutes
at about 95 "C, in 0.01 M citrate buffer (2.1 g citric acid
monohydrate/liter distilled water, pH adjusted to 6 with
NaOH). There were regular 1-minute intervals between
the heating periods, and at the end of the procedure the
specimens were left to cool down for more than 1 hour.I3
The sections were then incubated with anti-MIB-1 (Immunotech International, Marseilles, F r a n ~ e )or
~ antiPClO (Novocastra Laboratories, Newcastle-upon-Tyne,
England)5 monoclonal antibodies for either 2 hours at
room temperature or overnight at 4 "C, followed by incubation with biotinylated antimouse immunoglobulin G
for 30 minutes and with avidin-biotin complex reagent
(Vector Laboratories, Burlingame, CA) for another 30
minutes. The sections were subsequently reacted with
3,3'-diaminobenzidine tetrahydrochloride (DAB; Dojin
Chemicals, Tokyo, Japan) solution and 0.01% (weight/
volume) hydrogen peroxide for approximately 5 minutes
at room temperature, and counterstained with hematoxylin. Negative controls were obtained by substituting phosphate-buffered saline for the primary antibody.
Assessment of MIB-1 and PC10 Labeling Indices
The MIB-1 and PClO labeling indices are the percentages
of MIB-1- or PC10-positive cancer cells, respectively. The
immunostained sections were evaluated blindly to avoid
bias. Three randomly selected representative fields were
used to count the cancer cells to determine the indices.
A minimum of 1000 cancer cells (range, 1000 to 1500
cells) in every case was counted in 3 color photographs
(X350) to minimize variations originating from heteroge-
2282
CANCER June 1,1996 I Volume 77 / Number 11
FIGURE 1. Adenocarcinoma cells are positive for MIB-1 (a) and PClO (b), showing intranuclear staining (MIB-1 and PCIO irnmunostaining, x350).
neity within the sections. Cell counting was performed
by the same observer (a pathologist), who was blinded to
the identity of the patient from whom the specimen was
obtained. Reproducibility was determined by counting in
the same color photographs (twice in some cases) and
by counting in different color photographs taken from
other fields in the same cases. Variations between the
first counting and the second and variations among the
different fields were less than 5%.
Statistical Analysis
Statistical analysis was performed by the chi-square test
and Student’s t test, as appropriate, using a computerized
statistical package (HALBAU, Gendai Sugakusha, Tokyo,
Japan).
RESULTS
MIB-1 and PClO antigens were present in the nuclei of
cancer cells in both the adenocarcinomas (Figs. l a and
b) and the squamous cell carcinomas (Figs. 2a and b).6,8
The mean MIB-1 and PClO labeling indices before RT in
relation to the clinical stage and histologic subtype in the
14 patients with adenocarcinomas are shown in Table I,
whereas Table 2 shows these results for the 62 patients
with squamous cell carcinomas. There were no significant
relationships between stage or subtype and the two indices in either group of patients.
In patients with adenocarcinomas, the mean MIB- 1
labeling indices before RT and at 9 and 27 Gy were 28%,
21%, and 26%,respectively, and the mean PClO labeling
indices were 15%, 13%, and 14%, respectively (Table 3).
The mean MIB-1 and PClO labeling indices of the adenocarcinomas showed no remarkable change at 9 Gy or at
27 Gy (Figs. 3 and 4). In patients with squamous cell
carcinomas, the mean MIB- 1 labeling indices before RT
and at 9 and 27 Gy were 38%, 53%, and 26%, respectively,
and the mean PClO labeling indices were 23%, 23%, and
11%, respectively (Table 3). The mean MIB-1 labeling indices of the squamous cell carcinomas were significantly
increased at 9 Gy ( P < 0.01) and decreased at 27 Gy ( P
< 0.01; Fig. 3). The mean PClO index did not show any
remarkable change at 9 Gy and was significantly decreased at 27 Gy ( P < 0.01; Fig. 4). Table 3 demonstrates
significant differences in MIB-1 and PClO indices between adenocarcinomas and squamous cell carcinomas
before RT and at 9 Gy. However, at 27 Gy, the indices
for adenocarcinomas and squamous cell carcinomas had
similar values.
DISCUSSION
The present study showed that the mean MIB-1 and PClO
labeling indices of adenocarcinomas were significantly
lower than those of squamous cell carcinomas ( P < 0.001
and P < 0.001, respectively; Table 3). A similar cell cycle
trend was observed in pulmonary carcinomas. Gatter et
al.,I4in their investigation of 90 patients with pulmonary
carcinoma (27 with adenocarcinomas and 63 with squamous cell carcinomas), reported that the Ki-67 labeling
indices were lower in the adenocarcinomas than in the
squamous cell carcinomas. Kawai et al.lS reported that
the mean Ki-67 labeling index of pulmonary adenocarcinomas was 33%, whereas that of squamous cell carcinomas was 45%. Carey et
reported that the mean PCNA
labeling indexes of pulmonary adenocarcinomas and
squamous cell carcinomas were 19% and 32%, respectively. These findings, as well as those in the present
study, suggest that the proliferative activity of adenocarcinomas may be lower than that of squamous cell carcinomas, at least in terms of the percentage of the cycling cell
population.
The present study demonstrated differential changes
in the MIB-1 and PClO indices of tumor cells between
Response to Radiation in Cervical Carcinoma/Oka et al.
2283
FIGURE 2. Squarnous cell carcinoma cells are positive for MIB-1 (a) and PClO (b), showing intranuclear staining (MIB1 and PClO imrnunostaining
x 350).
TABLE 1
Mean MIB-1 and PClO Indices before Radiotherapy in 14 Patients
with Cervical Adenocarcinoma in Relation to the Clinical Stage and
Histologic Subtype
TABLE 2
Mean MIB-1 and PClO Indices before Radiotherapy in 62 Patients
with Cervical Squamous Cell Carcinoma in Relation to the Clinical
Stage and Histologic Subtype
Mean value pm SD (%b)
Mean value 2 SD (W)
Stage
2b
3b
3a
Subtype
EC
.4S
EM
'Total
n
MIB-1 index
PClO index
5
6
3
28 2 4
26 i ti
33 i ti
14 2 7
16 2 3
15 2 2
ti
ti
2
14
28 i ti
28 i 3
29 2 11
28 -C 6
14 i16 i15 I
15 5
4
7
2
5
iD: standard deviation: n: number of patients; EC: adenocarcinorna, endocenical type; EM: endometriadenocarcinoma; AS: adenosquamous carcinoma.
3id
adenocarcinomas and squamous cell carcinomas during
the period of RT. The MIB-1 labeling index of the squamous cell carcinomas significantly increased at 9 Gy and
then decreased to a lower level than before RT at 27 Gy.
However, the indices of the adenocarcinomas did not
show any remarkable increase during the period of RT. To
our knowledge, there has been no report to demonstrate
differences in cell cycle changes between adenocarcinomas and squamous cell carcinomas during RT. It is not
easy to explain this difference, because tumor cell kinetics
during RT are considered to be associated with such complicated biologic factors at the tissue level as reoxygenation, repopulation, regeneration, and recruitment. One
reason for the increase in the MIB-1 index of squamous
cell carcinomas at 9 Gy, however, appears to be the re-
Stage
1
2b
3b
4a
Subtype
LN
SN
K
Total
n
MIB-I index
PClO index
3
17
38
4
39 5 10
40 ? 14
37 i 14
43 i 17
35 ? 2
23 i 12
22 2 10
25 i 13
36
13
13
ti2
39 2 15
40 i 9
35 i 14
38 i 14
25 i 11
18 i 8
24 i I1
23 2 11
SD: standard deviation; n: number of patients: LN: large cell nonkeratinizing cancer; SN: small cell
nonkeratinizine cancer: K keratinizine cancer.
cruitment phenomenon, in which quiescent cells enter
the cell cycle via the changes in nutritional or oxygenating
circumstances induced by irradiation."'" Even if recruitment is one of the major associating factors in the early
radiation period, it would seem to be less responsible for
adenocarcinomas of the cervix. Opinions on the prognosis of cervical carcinoma in relation to histology are controversial. Some investigators have reported difference in
survival rates between patients with cervical adenocarcinomas and those with squamous cell carcinomas,"
whereas others have reported that patients with cervical
adenocarcinomas had a worse prognosis than those with
squamous cell carcinomas. We have already reported
that the 5-year survival rate of patients with Stage 111
2284
CANCER June 1, 1996 / Volume 77 I Number 11
TABLE 3
Mean MIB-1 and PClO Indices in Patients with Cervical Adenocarcinoma and Squamous Cell Carcinoma before and during Radiation Therapy
Mean values ? SD
(W
Index
Subtype
Before RT
9 GY
(n = 14)
(n = 9)
27 Gy
Ad
(n = 7 )
P < 0.001
MIB-1
P < 0.001
sq
(n = 62)
(n = 62)
Ad
in
1 5=2 513)
1
(n = 41)
(n = 8)
P < 0.001
PClO
P= 1
in
=
7)
P = 0.005
P = 0.451
sq
(n = 62)
(n = 62)
(n = 42)
S D standard deviation; before R T before radiotherapy; A d adenocarcinoma; Sq: squamous cell carcinoma; n: number of patients; Gy: Gray.
P< 0.01.n=41
40
P<O.Ol, n=62
T
60
1”
squamn-62
........0....
adeno n-14
p<O.Ol, n=41
squam,n-62
T T
3.1
.......
4-J”
.””
adeno, n-14
10p=o.12, n=9
0-
O i
I
p=O.55, n=7
I
I
1
I
1
1
0
9
18
27
Gy
Radiation dose
FIGURE 4. Mean P C I O index with standard deviation before and during
radiotherapy; 14 patients with adenocarcinornas ( . . . . .) and 62 patients
with squamous cell carcinomas (-).
cervical adenocarcinomas was 30%, poorer than that of
patients with squamous cell carcinomas, which have a
survival rate of 52%.”,” Similarly, the comparative radiation sensitivity of cervical adenocarcinomas and squamous cell carcinomas remains a subject of considerable
controversy. Some investigators have suggested that the
lower radiation sensitivity of adenocarcinomas explains
the poorer local control rate. However, others believe that
the poorer local control may be due to myometrial invasion rather than to lower radiation sensitivity.2o
Recently, we demonstrated that adenocarcinomas of
the cervix were rather radioresistant compared with squamous cell carcinomas when treated with high dose rate
intracavitary radiation.‘l The present results indicate that
the lower growth fraction in adenocarcinomas may be
one reason for the radiation resistance of this tumor, be-
cause a lower growth fraction indicates a large quantity
of Go phase cancer cells, which are regarded as radiation
resistant. The differences between adenocarcinomas and
squamous cell carcinomas in the MIB-1 and PClO indices
during RT may be associated with the differences in local
control, although the differences in intracellular radiation
sensitivity among various cell types are regarded as being
responsible for radiation sensitivity. Further investigation
of cell cycle kinetics during RT is required to elucidate
the precise mechanisms of tumor response to radiation
and to optimize the individualized application of RT.
REFERENCES
1.
Nielson IC, Smith RR, McLaren JR, Robrough JE. Carcinoma
of the uterine cervix: a study of 864 patients. Cancer
1967;20:86-92.
Response to Radiation in Cervical Carcinoma/Oka et al.
2.
3.
4.
5.
6.
7.
8.
9.
10.
11.
Yinen K, Nieminen V, Matti F, Widholm 0, Karjalainen 0.
Changing pattern of cervical carcinoma: a report of 709 cases
of invasive carcinoma treated in 1970- 1974. Gynecol Oncol
1985;20:378-86.
Gerdes J, Schwab U, Lemke H, Stein H. Production of a
mouse monoclonal antibody reactive with a human nuclear
antigen associated with cell proliferation. Int J Cancer
1983;31:13-20.
Key G, Becker MHG, Baron B, Duchrow M, Schluter C, Flad
H-D, et al. New Ki-67-equivalent murine monoclonal antibodies (MIB1-3) generated against bacterially expressed
parts of the Ki-67 cDNA containing three 62 base pair repetitive elements encoding for the Ki-67 epitope. Lab Invest
1993;68:629-36.
Waseem NH, Lane DP. Monoclonal antibody analysis of the
proliferating cell nuclear antigen (PCNA). Structural conversation and detection of a nucleolar form. J Cell Sci
1990;96:121-9.
Oka K, Hoshi T, Arai T. Prognostic significance of the PClO
index as a prospective assay for cervical cancer: treated with
radiation therapy alone. Cancer 1992;70: 1545-50.
Oka K, Nakano T, Arai T. Cytoplasmic p105 index is an accurate mitotic index, but is not related to prognosis in cenical
carcinoma. Arch Patkol Lab Med 1994; 118:506-9.
Oka K, Arai T. MIBl growth fraction is not related to prognosis in cervical carcinoma treated with radiotherapy. Znt J
Gynecol Pathol. 1996 1523-7.
Nakano T, Oka K. Transition of Ki-67 index of uterine cervcal tumors during radiation therapy. Cancer 1991;6851723.
Oka K, Nakano T, Hoshi T. Transient increases of growth
fraction during fractionated radiotherapy for cervical carcinoma: Ki-67 and PClO immunostaining. Cancer 1993;
722621 -7.
Arai T, Nakano T, Sakashita K, Nakamura YK, Fukuhisa K.
High-dose-rate remote afterloading intracavitary radiation
12.
13.
14.
15.
16.
17.
18.
19.
20.
21.
2285
therapy for cancer of the uterine cervix. A 20-year experience. Cancer 1992;69:175-80.
Landberg G, Roos G. Antibodies to proliferating cell nuclear
antigen as S-phase probes in flow cytometric cell cycle analysis. Cancer Res 1991;51:4570-4.
Shi SR, Key ME, Kalra K. Antigen retrieval in formalin-fixed,
paraffin-embedded tissues: an enhancement method for immunohistochemical staining based on microwave oven
heating of tissue sections. J Histochem Cytockem 1991;
39:741-8.
Gatter KC, Dunnill MS, Gerdes J, Stein H, Mason DY. New
approach to assessing lung tumors in man. J Clin Pathol
1986;39590-3.
Kawai T, Suzuki M, Kono S, et al. Proliferating cell nuclear
antigen and Ki-67 in lung carcinoma. Correlation with DNA
flow cytometric analysis. Cancer 1994;742468-75.
Carey FA, Fabbroni G, Lamb D. Expression of proliferating
cell nuclear antigen in lung cancer: a systemic study and
correlation with DNA ploidy. Histopathology 1992;20:499503.
Leminen A, Paavonen J, Forss M, Wahlstrom T, Vesterinen
E. Adenocarcinoma of the uterine cervix. Cancer 1990;65:539.
Eifel PJ, Morris M, Oswald MJ,Wharton JT, Delclos L. Adenocarcinoma of the uterine cervix. Prognosis and patterns of
failure in 367 cases. Cancer 1990;65:2507- 14.
Oka K, Nakano T, Arai T. Adenocarcinorna of the cervix
treated with radiation alone: prognostic significance of S100 protein and vimentin immunostaining. Obstet Gynecol
1992;79:347-50.
Fletcher GH, Rutledge FN, Delclos L. Adenocarcinoma of
the uterus. In: Fletcher GH, editor. Textbook of radiotherapy.
3rd edition. Philadelphia: Lea & Febiger, 1980:789-90.
Nakano T, Arai T, Morita S, Oka K. Radiation therapy alone
for adenocarcinoma of the uterine cervix. IntJ Radiat Oncol
Biol Pkys 1995;32:1331-6.
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