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Immunology |
Departments of Obstetrics and Gynecology [B-C. S., H-N. H., S-C. H.], Pathology [H-C. L., S-M. H.], and Graduate Institute of Immunology [H-N. H., S-M. H., R-H. L.], National Taiwan University College of Medicine, Taipei, Taiwan, 100
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ABSTRACT |
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 Top ABSTRACT INTRODUCTIONMATERIALS AND METHODSRESULTS AND DISCUSSIONREFERENCES |
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(IL-2R) on activated tumor-infiltrating
lymphocytes was down-regulated. Because IL-2R plays a pivotal
role in the development and propagation of functional T cells, its
depressed expression may result in poor function of tumor-reactive
cytotoxic lymphocytes. For elucidating the mechanism responsible for
down-regulation of IL-2R, a coculture model of in
vitro mixed autologous lymphocytes and tumor cells was
established. Kinetic analysis showed that cervical cancer cells
down-regulated IL-2R expression on encountered T cells. The amount
of IL-2R mRNA in tumor-infiltrating lymphocytes-derived
CD8+ T cells was compatible with that in the corresponding
activated CD8+ T cells. Additional evidence showed that
cervical cancer cells could induce the release of soluble IL-2R
expression on encountered T cells. By using protease inhibition assays
we demonstrated that tissue inhibitors of metalloproteinase abrogated
the cancer-mediated IL-2R proteolytic process and restored the
T-cell proliferation function. Immunohistochemical stainings further
revealed prominent metalloproteinase (MMP) expressions,
including MMP-1, MMP-2, and MMP-9, in cervical cancer tissues.
Additional in vitro studies showed that MMP-9 mediates
cleavage of IL-2R and down-regulates the proliferative capability of
cancer-encountered T cells. Our findings suggest a new role of MMPs in
tumor-mediated immunosuppression and provide a possible therapeutic
potential for patients with cervical cancer. |
INTRODUCTION |
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TopABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTS AND DISCUSSIONREFERENCES |
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3
on activated TILs was down-regulated (6, 7, 8, 9)
. Because
IL-2R plays a pivotal role in the development and propagation of
functional T cells, its depressed expression may result in poor
function of tumor-reactive cytotoxic lymphocytes.
Depressed immune responses have been observed in selected patients with
various types of tumors, including CC (10, 11, 12)
. Not
infrequently, cancer spread occurs despite the apparent presence of
TILs (12, 13, 14)
. In human CC as well as in other cancers,
tumor-specific autologous cytotoxic T cells are easily identified, but
the cytotoxic potential of freshly isolated TILs is usually not
expressed (10
, 13
, 14)
. This suggests the possible
existence of as yet unspecified and perhaps complex immunosuppressive
factors or cytokines that affect the antitumor capacity of these TILs
in the tumor microenvironment. Recently, we and others have
demonstrated that the expression of CD25 (IL-2R) on TILs derived
from various human cancers was down-regulated (6, 7, 8, 9)
.
Because IL-2R is an essential receptor for the proliferation of T
cells, its depressed expression in vivo may result in poor
clonogenicity of cytotoxic lymphocytes (15, 16, 17, 18)
and cause
immune suppression. Possible explanations for down-regulation of
IL-2R on TILs are as follows: (a) intrinsic defects of
IL-2R biosynthesis in the T cells of patients with cancer;
(b) inhibition of IL-2R synthesis in TILs by exogenous
mediators derived from cancer cells (7
, 9)
; and/or
(c) rapid cleavage of IL-2R by intrinsic or extrinsic
means within the tumor milieu.
In this study, we used a MLTC model to evaluate the kinetic expression
of IL-2R on in vitro-activated T cells and to infer the
mechanism responsible for down-regulation of IL-2R on TILs in human
CC. A coculture of MLNC from the same patient was used as control.
TILs, cancer cells, and noncancerous cells were isolated by a
mechanical dispersal technique as described previously (8
, 11)
. PBMCs from the same patient were used as the source of
autologous lymphocytes. We demonstrated for the first time that
tumor-derived metalloproteinases can induce the proteolytic cleavage of
IL-2R on activated T cells and suppress the proliferative capability
of cancer-encountered T cells.
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MATERIALS AND METHODS |
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TopABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTS AND DISCUSSIONREFERENCES |
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Autologous Coculture Assay.
PBMCs were partially activated with 2 µg/ml of PHA in a 37°C, 5%
CO2-humidified incubator for 2 h. The
activation and coculture procedures outlined were performed in
duplicate in 12 mm x 75 mm capped polystyrene test
tubes. In the experimental group (MLTC), 5 x 105 activated T cells were cultured with
1 x 106 autologous cancer cells
(1:2) in 4 ml of culture medium. In the control group (MLNC),
5 x 105 activated T cells were
cultured with 1 x 106 autologous
normal cervical stromal/epithelial cells. The immunophenotyping of
activated T cells in both groups was analyzed by flow cytometry on days
1, 3, and 5 of coculture. The supernatant was collected at the same
intervals and checked for the presence of sIL-2R by a standard ELISA
(Endogen, Inc., Woburn, MA).
Flow Cytometry Analysis.
Monoclonal antibodies labeled with FITC, PE, and PerCP
(Becton Dickinson Immunocytometry System; Becton Dickinson, Inc., San
Jose, CA) were used for three-color flow cytometry. The following
matchings were arranged: (a) anti-CD45-FITC + anti-CD14-PE (leucogate); (b) anti-CD3-FITC + anti-CD19-PE; (c) anti-CD3-FITC + anti-CD4-PE;
(d) anti-CD3-FITC + anti-CD8-PE; (e)
anti-CD4-FITC + anti-CD25-PE + anti-CD3-PerCP;
(f) anti-CD8-FITC + anti-CD25-PE + anti-CD3-PerCP; (g) anti-CD4-FITC + anti-HLA-DR-PE + anti-CD3-PerCP; and (h)
anti-CD8-FITC + anti-HLA-DR-PE + anti-CD3-PerCP. A
Simultest control (mouse IgG1-FITC + IgG2a-PE) was used as
background control. Three-color flow cytometry was performed on a
FACScalibur (Becton Dickinson, Inc.). Leukogate was used to
measure the proportion of lymphocytes in the sample being studied
without any scatter gates.
Purification of CD8+ CTLs.
An indirect magnetic labeling system with a MACS (Miltenyi Biotec,
Gladbach, Germany) was used for purification of
CD8+ T cells. In brief, the cell suspension was
incubated in a cold room (6°C to 12°C) for 10 min with 20 µl of
Hepten-Antibody Cocktail added (containing CD4, CD11b, CD16, CD19,
CD36, and CD56 antibodies; Miltenyi Biotec). After repeated washing, 20
µl of MACS anti-Hepten microbeads/107 total
cells were added. The magnetically labeled cells were passed through a
MACS separator in the magnetic field. The effluent was collected as a
negative fraction representing the enriched
CD8+ T-cell fraction. The cell purity was checked
by flow cytometry after labeling with anti-CD8-FITC.
Competitive RT-PCR Analysis.
Total cellular RNA of MACS-purified CD8+ T cells
was extracted and reverse-transcribed (0.1–0.5 mg of RNA) in the
presence of IL-2R oligonucleotide primers (Clontech Laboratories,
Inc., Palo Alto, CA). First-strand cDNA was synthesized in a DNA
thermal cycler (GeneAmp DNA thermal cycler 480; Perkin-Elmer Corp.,
Norwalk, CT), and the resulting cDNA was amplified by PCR. For
competitive analysis of IL-2R, another set of synthetic competitors
(IL-2R mimic primers) with a different length for amplification was
used (Clontech Laboratories Inc.). Human ß-actin primers and
ß-actin mimic primers were used as positive controls.
Preparation of Protease Inhibitors.
Protease inhibitors (Boehringer Mannheim Biochemicals GmbH, Mannnheim,
Germany, and Calbiochem, La Jolla, CA) for a broad spectrum of
inhibition were prepared. The selected protease inhibitors included
antipain-dihydrochloride (papain, trypsin, cathepsin A- and
B-inhibitor), bestatin (amino peptidase inhibitor), chymostatin
(-, ß-, 
-, and 
-chymotrypsin-inhibitor), E-64 (cysteine
protease inhibitor), leupeptin (serine and cysteine protease, plasmin,
trypsin, papain, and cathepsin B-inhibitor), pepstatin (aspartate
protease), aprotinin (serine protease inhibitor, specific for plasmin,
kallikrein, trypsin, and chymotrypsin), TIMP-2
(Mr 72,000 and
Mr 92,000 gelatinases
inhibitor), and MMP-inhibitor I (a broad spectrum gelatinase
inhibitor). All reagents were prepared in azide-free deionized
H2O. Titrated concentrations of individual
protease inhibitors were prepared according to the recommended working
formula of the reagent supplier (Boehringer Mannheim Biochemicals).
Drug toxicity was tested by the addition of titrated concentrations of
reagents in lymphocyte culture for 24 h with >90% viable cells
by the trypan-blue staining method.
Protease Inhibition Assay.
Different protease inhibitors were added to the established MLTC. For
maximal expression of IL-2R on the surface of T cells, PBMCs were
fully activated with 10 µg/ml PHA for 24 h before MLTC.
Autologous cancer cells (1 x 106)
were first cultured in 1 ml of culture medium containing protease
inhibitors. The final concentrations of protease inhibitors as
recommended in each tube were as follows: (a)
antipain-dihydrochloride, 5 and 50 µg/ml; (b) aprotinin, 1
and 10 µg/ml; (c) bestatin, 4 and 40 µg/ml;
(d) chymostatin, 6 and 60 µg/ml; (e) E-64, 1
and 10 µg/ml; (f) leupeptin, 1 and 10 µg/ml;
(g) pepstatin, 1 and 10 µg/ml; (h) TIMP-2, 0.1
and 1 µg/ml; and (i) MMP-inhibitor I, 10 and 100
µM/ml (Calbiochem). Constant numbers
(5 x 105 cells) of PHA-activated
T cells were then added to the MLTC. In the control group, which
included only activated T cells and coculture of activated T cells with
cancer cells, no protease inhibitor was added. The supernatant of each
culture tube was collected on the third day of coculture, and the
concentration of sIL-2R was checked by ELISA.
Effect of MMP-inhibitors on T-cell Proliferation.
PBMCs were fully activated with 10 µg/ml PHA for 24 h before
MLTC. Autologous cancer cells (1 x 106) were first cultured in 1 ml of culture
medium containing 1 µg/ml TIMP-2 or 100 µM/ml
MMP-inhibitor I for 2 h and cocultured with PHA-activated T cells
(5 x 105 cells). In the control
group, no TIMP-2 was added. Two days after MLTC, T cells were purified
by MACS as described previously and cultured at constant numbers
(2 x 105 cells/well) in the
presence of titrated IL-2 concentrations (1 IU/ml, 3 IU/ml, and 10
IU/ml). Titrated thymidine ([3
H]TdR) was added
during the last 18 h of culture. Cells were harvested, and
thymidine incorporation was counted by liquid scintillation. All assays
were performed in triplicate.
Expression of MMPs (3)
, MT-MMPs, and TIMPs in CC
Tissue.
An avidin-biotin-peroxidase complex immunohistochemical staining method
was performed for examination of the expression patterns of MMP-1, -2,
-3, -7, -8, -9, -13, MT1-MMP, MT2-MMP, TIMP-1, TIMP-2, and TIMP-3 in
formalin-fixed and paraffin-embedded tissue sections. The antibodies
were obtained from Chemicon, Inc. (Temecula, CA), and their
specificities were provided by the manufacturer.
Analysis of MMP-9 Activity by Gelatin Zymography.
Tumor cell nests were carefully separated and scrapped from underlying
stromas by a microdissection method. The scrapped tissues were
homogenized with lysis buffer containing glycerol (10%), Triton X-100
(1%), sodium PPi (1 mM), NaCl (137
mM), EDTA (5 mM), sodium orthovanadate (1
mM), NaF (10 mM), and Tris (pH 7.9; 20
mM). T cells isolated from PBMCs were activated and
cultured as described previously.
The gelatinolytic activity of MMP-9 was determined by gelatin-substrate gel electrophoresis. Aliquots of supernatants of coculture or cell lysates (15 µg protein/lane) were applied, without heating or reducing, to a 10% SDS-polyacrylamide gel containing 1 mg/ml gelatin. After electrophoresis, gels were washed for 1 h at room temperature in a 2% Triton X-100 solution before being transferred to a reaction buffer containing 50 mM Tris-HCl (pH 7.4) 0.15 M NaCl, and 10 mM CaCl2 and incubated at 37°C overnight. The gel was stained with 0.1% Coomassie Blue in 50% methanol:10% acetic acid and destained in 20% methanol:10% acetic acid. Clear zones of gelatin lysis against a blue background stain indicated the presence of enzyme. Quantitative analysis of gelatinolytic activity was achieved by scanning densitometry of the zymograms (IS-1000 Digital Imaging System; Alpha Innotech Corp., San Leandro, CA).
Effects of Recombinant MMPs on the Function of Activated T Cells.
PBMCs were fully activated with 10 µg/ml PHA for 24 h.
Recombinant MMP-1, MMP-2, and MMP-9 proteases (Calbiochem, Inc.,
Cambridge, MA) were added to the cultures at a final concentration of 1
µg/ml, and the PBMCs were incubated for an additional 6 h. The
activated cells were labeled with anti-CD25-PE and checked by flow
cytometry (8
, 11)
. In addition, MMP-treated T cells were
purified and cultured at constant numbers (2 x 105 cells/well) in the presence of titrated IL-2
concentrations (1 IU/ml, 3 IU/ml, 10 IU/ml, and 100 IU/ml), and the
T-cell proliferation assays were conducted as described above. Studies
were performed in triplicate.
Statistical Analysis of Data.
All results were expressed as mean ± SE unless stated
otherwise. Data analysis was performed with SAS (Statistical Analysis
System, R.6.12; SAS Institute, Inc., Cary, NC). Kinetic data were
analyzed by a generalized estimation equation for correlated data of
repeated measurements. Statistical significance was defined by a
P of <0.05.
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RESULTS AND DISCUSSION |
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TopABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTS AND DISCUSSIONREFERENCES |
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Expression on Encountered T Cells.
and HLA-DR on activated T cells was measured by flow cytometry
(8
, 11)
. The kinetic expression of IL-2R on activated T
cells in the MLNC was compatible with the known physiological pattern
(16
, 17)
, being highest on day 3 and having
progressively decreased by day 5. Thus, the noncancerous cells had no
influence on the level of IL-2R expression in MLNC. In contrast, CC
cells seemed to affect the expression of IL-2R by T cells in MLTC.
On day 1 post-coculture, the MFI of IL-2R expression on activated
CD4+ cells was similar in both MLTC and MLNC. It
was noteworthy that the MFI of IL-2R on CD4+
cells was significantly lower in MLTC than in MLNC on day 3
(170.7 ± 17.0 versus 360.1 ± 14.7) and day 5 (94.1 ± 6.1 versus
174.3 ± 23.6). A similar finding was obtained with
CD8+ cells (139.0 ± 8.9
versus 340.0 ± 28.9 on day 3 and
64.4 ± 10.7 versus 196.3 ± 28.7 on day 5 for MLTC and MLNC, respectively; Fig. 1, A and B
). However, there was no difference in
the kinetic expression patterns of HLA-DR on activated T cells
(CD4+ or CD8+) between MLTC
and MLNC (Fig. 1, C and D)
. The expression of
HLA-DR on activated T cells of MLTC and MLNC was low on day 1 and
elevated on day 3, and remained persistently high on day 5. Our
previous study had shown that the expression of HLA-DR, and CD69 as
well, on TILs isolated from patients with CC was not altered
(8)
. Together, these findings indicate that CC cells can reduce IL-2R expression on encountered T cells in a
specific manner.
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on TILs Is Not Mediated at the
Transcriptional Level. on TILs
in CC was mediated at the transcriptional level.
CD8+ T cells were isolated with a purity of
>97% from TILs, unstimulated PBMCs, and PHA-activated PBMCs.
Expression of IL-2R mRNA was measured by competitive RT-PCR. The
amount of IL-2R mRNA in TIL-derived CD8+ T
cells was comparable with that in the activated
CD8+ T cells. In both TILs and activated
CD8+ T cells, the amount of IL-2R mRNA was
about 100-fold of that expressed in nonstimulated PBMC-derived
CD8+ cells (Fig. 1, E and F)
. A deficiency in IL-2 protein and IL-2R expression
despite adequate levels of IL-2 mRNA has also been shown in TILs from
patients with breast cancer (7)
. Because the IL-2R mRNA
was abundant in TILs, cancer-mediated suppression did not occur at the
transcriptional level.
CC Cells Enhance the Release of sIL-2R from Activated T Cells.
The possibility exists that the CC-associated decreased expression of
IL-2R results from excessive shedding of IL-2R from T cells. To
clarify this possibility, we analyzed the amounts of sIL-2R in the
supernatants of MLTC and MLNC. The mean concentration of sIL-2R in
MLNC supernatants was 1045.1 ± 17.3 pg/ml and increased
about 2- and 3-fold by day 3 and day 5, respectively. The cumulative
sIL-2R concentration in the supernatants of MLNC was compatible with
the pattern of natural shedding of IL-2R (16
, 17)
.
However, the mean concentration of sIL-2R in the supernatants of
MLTC was significantly higher than that in MLNC in day-3 and day-5
cocultures (increased more than 3- and 4-fold, respectively;
P < 0.001; Fig. 2A
). It became evident that CC cells could enhance the release
into the surroundings of sIL-2R from activated T cells, and that a
proteolytic cleavage might mediate the process (17
, 18)
.
|
.. For achieving maximal expression of IL-2R, PBMCs were
fully activated with 10 µg/ml PHA for 24 h before MLTC. After 3
days of coculture, the concentration of sIL-2R in the MLTC group was
3-fold that in the T-cells-only group (6001.0 ± 679.6 pg/ml versus 2241.3 ± 197.3 pg/ml;
n = 6, P < 0.001).
Protease inhibitors such as E-64, aprotinin, bestatin, leupeptin,
pepstatin, chymostatin, and antipain-dihydrochloride had no apparent
activity in suppressing CC cell-mediated proteolytic cleavage of
IL-2R (Fig. 2B)
.
TIMP-2 and MMP-inhibitor I significantly inhibited the CC cell-induced
IL-2R cleavage in a dose-dependent pattern (Fig. 2B)
. At
a concentration of 1 µg/ml of TIMP-2, cancer-induced IL-2R
cleavage was totally inhibited. The concentration of sIL-2R in the
TIMP-2-MLTC group was 1953.1 ± 81.2 pg/ml, compared
with a concentration of 6001.0 ± 679.6 pg/ml in the
MLTC group without inhibitor (n = 6;
P = 0.004). At a concentration of 0.1 µg/ml
TIMP-2, partial inhibition existed (3136.2 ± 327.7
pg/ml versus 6001.0 ± 679.6 pg/ml;
P = 0.011). At a concentration of 100
µM/ml MMP-inhibitor I, IL-2R cleavage was
also totally inhibited. The concentration of sIL-2R in the
MMP-inhibitor I-MLTC group was 2114.7 ± 165.7 pg/ml,
compared with a concentration of 6660.1 ± 640.5 pg/ml
in the MLTC group without inhibitor (n = 6;
P = 0.001). At a concentration of 10
µM/ml of MMP-inhibitor I, partial inhibition
existed (3264.2 ± 226.2 pg/ml versus
6660.1 ± 640.5 pg/ml; P = 0.006). The
finding indicates that an MMP-mediated proteolytic process is likely to
be responsible for the IL-2R cleavage, and that MMP inhibitors block
this process.
MMP Inhibitors Restore IL-2-promoted Proliferation Function of
TILs.
To examine whether TIMP-2 or MMP-inhibitor I added to cultures could
restore the proliferative function of cancer-encountered T cells, we
also performed an IL-2-promoted T-cell proliferation assay. In the MLTC
experiments illustrated in Fig. 2C
, cancerencountered T
cells proliferated poorly in the presence of IL-2. However, at a
concentration of 1 µg/ml TIMP-2 or 100 µM/ml
MMP-inhibitor I, the proliferative ability of cancer-encountered T
cells could be restored in an IL-2 dose-dependent manner, which
indicated that MMP inhibitors are functionally capable of restoration
of the T-cell proliferation function by blocking the cancer-induced
IL-2R cleavage.
MMP-1, -2, and -9 Are Abundantly Expressed by CC Cells.
We then studied the expression of various types of MMPs, including
MMP-1, -2, -3, -7, -8, -9, and -13, two membrane-type MMPs (MT1 and
MT2), and three types of inhibitors (TIMP-1, -2, and -3) by
immunohistochemical staining in 30 cases of CC tissues. MMP-1, MMP-2,
and MMP-9 are abundantly expressed in most, if not all cancer cells,
but are not expressed or are only very weakly expressed in normal
cervical epithelial or stromal cells (Fig. 3, A
–D, data not shown). The expression of other
types of MMPs and two types of membranous MMPs was not detected or was
only weakly expressed. Diffusely weak stainings of TIMP-1, TIMP-2, and
TIMP-3 were observed in selected cases of CC (weakly expressed in
10–20% of cases). Low TIMP expression and/or increased MMP:TIMP ratio
in tissues were reported to associate with a poor prognosis for CC
(19
, 20) . In the gelatin zymographic study (Fig. 3E)
, the gelantinolytic activities of MMP-9
(Mr 92,000) were strongly expressed in
all CC cells but varied widely in supernatants of MLTC (data not
shown). The enzyme activities of MMP-9 in activated T cells were about
1–8% of CC cells.
|
. cleavage, we
directly incubated activated T cells with 1 µg/ml recombinant MMP-1,
MMP-2, or MMP-9 protease. As demonstrated by flow-cytometric analysis,
MMP-1 had no effect on the expression of IL-2R on activated T cells
(Fig. 4A)
. MMP-9 and, to a lesser extent, MMP-2, can down-regulate
the expression of IL-2R on activated T cells (Fig. 4, B and C)
. In the experiments illustrated in Fig. 4D
, MMP-2-treated T cells proliferated well in an IL-2
dose-dependent manner, which was compatible with the pattern of
activated T-cell controls. MMP-9-treated T cells proliferated poorly in
the presence of low-dose IL-2, but their proliferative ability could be
restored by either IL-2 at a higher dose or MMP-inhibitors. Thus, the
observed cancer-mediated IL-2R down-regulation can be attributed
mainly to proteolytic cleavage by MMP-9 from cancer cells. Because
IL-2R plays a pivotal role in the development and propagation of
functional T cells, cleavage of IL-2R down-regulates the
proliferative capability of T cells (16
, 17
; Figs. 2C
and 4D
).
|
MMPs are abundant in a variety of human cancer cells, including those
of CC (19
, 20
, 25)
. Cancer-derived MMPs may trigger the
proteolytic cleavage of cytokines and their receptors, including tumor
necrosis factor-R (28
, 29)
, IL-6R (29
, 30)
, and also IL-2R, as shown in the present study. A recent
study by Wang et al. (31)
showing that
MMP-9-deficient mice cannot resolve a contact hypersensitivity reaction
further implies the down-regulatory role of MMP-9 in immune responses.
Moreover, cancer-encountered T cells lacking IL-2R signaling may be
resistant to Fas-mediated apoptosis (32
, 33)
, which may
reciprocally explain the abundance of, but the relatively poor function
of, TILs within the tumor milieu.
In conclusion, the mechanism for apparent inhibition of IL-2R
expression on activated T cells in tumors is governed by a cancer
cell-directed and MMP-dependent cleavage. Cancer-derived MMP-9, and
other MMPs as well, can inhibit the proliferative function of
tumor-encountered T cells, and this may explain the relative anergic
state of TILs. Through the MMP-proteolytic pathway, cancer cells may
facilitate immunoescape, cancer invasion, and metastasis. It is
noteworthy that TIMP activity was significantly lower in CC tissues
than in other gynecological cancer tissues (20)
, and that
a reduction in TIMP expression in tissue is associated with a poor
prognosis for CC (19)
. Reversal of tumor-induced
immunosuppression has been a major concern regarding prospects for
adoptive T-cell therapy. Our finding should provide important insights
into the understanding of the interaction between cancer cells and the
immune system, as well as into the development of anticancer
strategies.
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FOOTNOTES |
|---|
1 This work was supported by grants from the
National Science Council (NSC88-2314-B-002-382, NSC89-2314-B002-162,
and NSC89-2314-B002-294) and the National Health Research Institute
(NHRI-GT-EX89S704 L), and research grants from the National Taiwan
University Hospital (NTUH.88A016, NTUH.89A011, and NTUH89-S1537). 
2 To whom requests for reprints should be
addressed, at R-H. L., AbGenomics Co., 2F, No.32 Lane 358, Juikuang
Road, Neihu, Taipei, Taiwan. Fax: 886-2-2627-2708; E-mail: rhlin{at}abgenomics.com S-C. H., at Department of Obstetrics and
3 The abbreviations used are: IL-2R,
interleukin-2 receptor ; CC, cervical cancer; TILs,
tumor-infiltrating lymphocytes; MLTC, mixed autologous lymphocyte and
tumor-cell coculture; MLNC, mixed autologous lymphocytes and
non-cancerous stromal/epithelial cells coculture; PBMC, peripheral
blood mononuclear cell; PHA, phytohemagglutinin; PE, phycoerythin;
PerCP, peridinin chlorophyll protein; MACS, magnetic activated cell
sorter; TIMP, tissue inhibitor of metalloproteinase; RT-PCR, reverse
transcription-PCR; MFI, mean fluorescence intensity; sIL-2R, soluble
form of IL-2R; MMPs, matrix metalloproteinases.
Received 2/17/00. Accepted 10/26/00.
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REFERENCES |
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TopABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTS AND DISCUSSIONREFERENCES |
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): physiology of cell surface shedding and renal catabolism, and suppression of catabolism by antibody binding. J. Exp. Med., 183: 1587-1602, 1996. subunit of the human T cell interleukin 2 receptor, by Der p 1, a major mite allergen with cysteine protease activity. J. Exp. Med., 187: 271-275, 1998. precursor by metalloproteinases. Nature (Lond.), 370: 555-557, 1994.[Medline]
processing by a metalloproteinase inhibitor. Nature (Lond.), 370: 558-561, 1994.[Medline]
)-deficient T cells expressing a transgenic antigen receptor. J. Immunol., 15: 3738-3745, 1997.
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 C. Cheung, D. Marchant, E. K.-Y. Walker, Z. Luo, J. Zhang, B. Yanagawa, M. Rahmani, J. Cox, C. Overall, R. M. Senior, et al. Ablation of Matrix Metalloproteinase-9 Increases Severity of Viral Myocarditis in Mice Circulation, March 25, 2008; 117(12): 1574 - 1582. [Abstract] [Full Text] [PDF]
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 P. Van Lint and C. Libert Chemokine and cytokine processing by matrix metalloproteinases and its effect on leukocyte migration and inflammation J. Leukoc. Biol., December 1, 2007; 82(6): 1375 - 1381. [Abstract] [Full Text] [PDF]
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 D.-W. Liu, Y.-C. Yang, H.-F. Lin, M.-F. Lin, Y.-W. Cheng, C.-C. Chu, Y.-P. Tsao, and S.-L. Chen Cytotoxic T-Lymphocyte Responses to Human Papillomavirus Type 16 E5 and E7 Proteins and HLA-A*0201-Restricted T-Cell Peptides in Cervical Cancer Patients J. Virol., March 15, 2007; 81(6): 2869 - 2879. [Abstract] [Full Text] [PDF]
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N. L. Webster and S. M. Crowe Matrix metalloproteinases, their production by monocytes and macrophages and their potential role in HIV-related diseases J. Leukoc. Biol., November 1, 2006; 80(5): 1052 - 1066. [Abstract] [Full Text] [PDF]
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 M. Stefanidakis and E. Koivunen Cell-surface association between matrix metalloproteinases and integrins: role of the complexes in leukocyte migration and cancer progression Blood, September 1, 2006; 108(5): 1441 - 1450. [Abstract] [Full Text] [PDF]
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P. E. Van den Steen, I. Van Aelst, V. Hvidberg, H. Piccard, P. Fiten, C. Jacobsen, S. K. Moestrup, S. Fry, L. Royle, M. R. Wormald, et al. The Hemopexin and O-Glycosylated Domains Tune Gelatinase B/MMP-9 Bioavailability via Inhibition and Binding to Cargo Receptors J. Biol. Chem., July 7, 2006; 281(27): 18626 - 18637. [Abstract] [Full Text] [PDF]
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K. J. Garton, P. J. Gough, and E. W. Raines Emerging roles for ectodomain shedding in the regulation of inflammatory responses J. Leukoc. Biol., June 1, 2006; 79(6): 1105 - 1116. [Abstract] [Full Text] [PDF]
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E. Mortier, A. Quemener, P. Vusio, I. Lorenzen, Y. Boublik, J. Grotzinger, A. Plet, and Y. Jacques Soluble Interleukin-15 Receptor {alpha} (IL-15R{alpha})-sushi as a Selective and Potent Agonist of IL-15 Action through IL-15Rbeta/{gamma}: HYPERAGONIST IL-15{middle dot}IL-15R{alpha} FUSION PROTEINS J. Biol. Chem., January 20, 2006; 281(3): 1612 - 1619. [Abstract] [Full Text] [PDF]
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 Z. Hu, X. Huo, D. Lu, J. Qian, J. Zhou, Y. Chen, L. Xu, H. Ma, J. Zhu, Q. Wei, et al. Functional Polymorphisms of Matrix Metalloproteinase-9 Are Associated with Risk of Occurrence and Metastasis of Lung Cancer Clin. Cancer Res., August 1, 2005; 11(15): 5433 - 5439. [Abstract] [Full Text] [PDF]
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 E. Godefroy, A. Moreau-Aubry, E. Diez, B. Dreno, F. Jotereau, and Y. Guilloux {alpha}v{beta}3-dependent cross-presentation of matrix metalloproteinase-2 by melanoma cells gives rise to a new tumor antigen J. Exp. Med., July 5, 2005; 202(1): 61 - 72. [Abstract] [Full Text] [PDF]
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 B.-C. Sheu, S.-H. Chiou, H.-H. Lin, S.-N. Chow, S.-C. Huang, H.-N. Ho, and S.-M. Hsu Up-regulation of Inhibitory Natural Killer Receptors CD94/NKG2A with Suppressed Intracellular Perforin Expression of Tumor-Infiltrating CD8+ T Lymphocytes in Human Cervical Carcinoma Cancer Res., April 1, 2005; 65(7): 2921 - 2929. [Abstract] [Full Text] [PDF]
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 E. Mortier, J. Bernard, A. Plet, and Y. Jacques Natural, Proteolytic Release of a Soluble Form of Human IL-15 Receptor {alpha}-Chain That Behaves as a Specific, High Affinity IL-15 Antagonist J. Immunol., August 1, 2004; 173(3): 1681 - 1688. [Abstract] [Full Text] [PDF]
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S. J. McMillan, J. Kearley, J. D. Campbell, X.-W. Zhu, K. Y. Larbi, J. M. Shipley, R. M. Senior, S. Nourshargh, and C. M. Lloyd Matrix Metalloproteinase-9 Deficiency Results in Enhanced Allergen-Induced Airway Inflammation J. Immunol., February 15, 2004; 172(4): 2586 - 2594. [Abstract] [Full Text] [PDF]
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B.-C. Sheu, H.-C. Lien, H.-N. Ho, H.-H. Lin, S.-N. Chow, S.-C. Huang, and S.-M. Hsu Increased Expression and Activation of Gelatinolytic Matrix Metalloproteinases Is Associated with the Progression and Recurrence of Human Cervical Cancer Cancer Res., October 1, 2003; 63(19): 6537 - 6542. [Abstract] [Full Text] [PDF]
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 R. Visse and H. Nagase Matrix Metalloproteinases and Tissue Inhibitors of Metalloproteinases: Structure, Function, and Biochemistry Circ. Res., May 2, 2003; 92(8): 827 - 839. [Abstract] [Full Text] [PDF]
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 K.-H. Chao, M.-Y. Wu, J.-H. Yang, S.-U. Chen, Y.-S. Yang, and H.-N. Ho Expression of the interleukin-2 receptor {alpha} (CD25) is selectively decreased on decidual CD4+ and CD8+ T lymphocytes in normal pregnancies Mol. Hum. Reprod., July 1, 2002; 8(7): 667 - 673. [Abstract] [Full Text] [PDF]
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B.-C. Sheu, R.-H. Lin, H.-C. Lien, H.-N. Ho, S.-M. Hsu, and S.-C. Huang Predominant Th2/Tc2 Polarity of Tumor-Infiltrating Lymphocytes in Human Cervical Cancer J. Immunol., September 1, 2001; 167(5): 2972 - 2978. [Abstract] [Full Text] [PDF]
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, MLNC). The data are representative
of six independent experiments of cocultures in triplicate.
A, MFI of IL-2R
), and MLTC in
the presence of MMP-inhibitors (•). The IL-2-dependent
[3H]TdR proliferation was analyzed. Data from six
independent experiments are presented as mean ± SE.
). Data
from four independent experiments are presented as mean ± SE.