复旦大学
论文集（2007）
Taxotere-Cisplatin Chemotherapy in Lung Cancer Patients
Preferentially Depletes Regulatory T Cells and Expands
CD44highCD62Llow T Cells Before Their Subsequent Recovery
to Original Proportions
上海医学院 04 级 温锦娣
上海医学院免疫学系 储以微 副教授
Abstract:
Purpose: The immunosuppressive actions of chemotherapy, chiefly its lymphodepleting effects,
had long been regarded as a hindrance for active immunity induction in the treatment of malignant
cancer. Recently chemotherapy has, however, been shown to benefit anti-tumor immunity when
combined with various cancer immunotherapies. To address the mechanism behind this seemingly
paradoxical phenomenon, we investigated for the first time the change in the immune pattern at
various time points in cancer patients undergoing chemotherapy, focusing on regulatory T cells
and effector memory T cells.
Experimental Design: The frequencies of various lymphocyte subsets and expression of FOXP3 and
intracellular IFN- in the peripheral blood of 23 non-small cell lung cancer (NSCLC) patients
following taxotere-cisplatin chemotherapy were dynamically analyzed at day -1, day 3 to 5 and
day 7 to 9 by three-color flow cytometry and intracellular CD8+IFN- cytokine detection
respectively.
Results: Compared to other lymphocyte subpopulations, CD4+CD25+FOXP3+ regulatory T cells were
selectively reduced at day 3 to 5, and subsequently recovered to its original frequency at day
high
low
7 to 9. Notably, the frequency of CD44 CD62L effector memory T cells peaked at day 3 to 5.
The level of intracellular CD8+IFN- secretion increased significantly at day 3 to 5 and
maintained at a relatively high level throughout day 7 to 9.
Conclusions: Chemotherapy induces a dynamic change of immune pattern in cancer patients. In vivo
immune environment between day 3 and 5 after taxotere-cisplatin chemotherapy, when Tregs are
selectively reduced and effector memory T cells undergo significant proliferation, before their
subsequent restoration to pre-chemotherapeutic levels, might be the key to devising successful
tumor immunotherapy strategies.
Key words: regulatory T cells, effector memory T cells, chemotherapy, lymphodepletion, immune
pattern
Introduction
Chemotherapy has remained as the mainstay of anticancer therapy due to its direct
tumoricidal properties. Its adverse immunosuppressive effects had been considered
unfavorable for eliciting anti-tumor immunity and were therefore merely tolerated.
The increasing use of immunotherapies in clinical settings has, however, driven
researchers to combine well-established chemotherapeutic regimens with novel
therapies such as tumor vaccination and adoptive cell transfer therapy. Surprisingly
chemotherapy has, in certain cases, markedly augmented antitumor immunity induced
by cancer immunotherapies both in animal models (1-3) and cancer patients (4-6), and
such combined therapy has become a very promising therapeutic approach for malignant
neoplasms, compared with the sole use of either treatment. Consequently the above
perspective that chemotherapy does not favor the induction of effective anti-tumor
immunity was revolutionized.
The team led by Rosenberg, who successfully achieved objective clinical response
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Dynamic change of immune pattern induced by chemotherapy
in 51% of patients with metastatic melanoma receiving adoptive cell transfer therapy
following chemotherapy (7), proposed that the enhanced efficacy of anti-tumor
immunity after lymphodepleting chemotherapy may be attributed to the abrogation of
tumor immunotolerance by mechanisms still being poorly elucidated (8).
Among them a key player is regulatory T cell (Tregs), the phenotypic marker of
which includes CD4, CD25(IL-2R) (9), GITR, CTLA-4, and most importantly, FOXP3 (10).
Treg plays a significant role in maintaining immunotolerance to self/tumor antigens
(11, 12) by suppressing the functions of self/tumor-reactive CD4+CD25- and CD8+ T
cells (13); and therefore its depletion can lead to autoimmune diseases (14, 15),
but at the same time enhance anti-tumor immunity (16, 17). The frequency of Tregs
in the peripheral blood of cancer patients is elevated (18-20) and increases with
malignancy (21, 22). Remarkably, its frequency and suppressive functions have been
shown to be preferentially reduced following chemotherapy in a few animal models (15,
23, 24). Such down-regulation of Treg proportion and functions has been proposed to
be a significant reason for the success of post-chemotherapeutic cancer
immunotherapies (25, 26).
The lymphopenic state induced by chemotherapy may also stimulate the preferential
homeostatic proliferation of residue weak self/tumor-reactive naïve T cells in the
lymphopenic state, which are skewed towards an effector memory T cell (TEM) phenotype
(CD44highCD62Llow) and function (27, 28), thereby shaping the T lymphocyte repertoire
towards effective reactivity to tumor antigens.
Nevertheless, the altered immune pattern induced by chemotherapy, especially the
question as to which subpopulations undergo changes and how do these changes vary
with time following chemotherapy has thus far remained unanswered. Therefore at day
-1, day 3 to 5 and day 7 to 9 after chemotherapy, through stepwise analysis of the
proportional change of each lymphocyte subpopulation in the peripheral blood, we
examined the dynamic change of Treg frequencies after chemotherapy in cancer patients.
We focused on whether it was selectively depleted by chemotherapy compared to other
lymphocyte subpopulations, and to target the time-point at which Treg frequency is
significantly reduced before recovering to its original state. This study also aims
to analyze the variation of effector memory T cell frequencies with respect to
different time points after chemotherapy, which is highly dynamic (29), during
reconstitution of the host’s immune system. Further, we also performed intracellular
staining of IFN- secretion in CD8+ T cells as a functional detection of the
immunologic state of chemotherapy-treated patients at these time points.
Materials and Methods
Patients and normal donors. Heparinized peripheral blood samples were collected
from 23 patients (19 men, 4 women, mean age 62, and range 41-76) with inoperable
non-small cell lung cancer. Most of the patients had either Stage IIIA (n=3), Stage
IIIB (n=6) or Stage IV tumor (n=14). The patients were undergoing standard
taxotere-cisplatin chemotherapeutic regimen at the time of study. Having obtained
informed consent from each individual according to the Declaration of Helsinki,
samples of 4 ml peripheral blood were obtained at different time points: on the day
before immunosuppressive chemotherapeutic drugs were administered (day -1), and day
3~4, day 7~8 after chemotherapy. Peripheral blood mononuclear cells (PBMCs) were
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obtained by the centrifugation of whole blood on a Ficoll/Hypaque gradient (Sinopharm
Chemical Reagent, China). PBMCs from 10 normal adults served as controls.
Information regarding patient history and tumor stage was recorded. Staging was
performed according to the AJCC/UICC classification for non-small cell lung cancer.
Flow Cytometric Analysis. PBMCs were stained for cell surface molecules and
intracellular markers (FOXP3, IFN-) to determine their immunophenotype with
monoclonal fluorescein isothiocyanate (FITC)-conjugated anti-CD3 (Sigma-Aldrich, MO,
USA), FITC-conjugated anti-CD4 (GeneMay, CA, USA), phycoerythrin-cyanine 5
(PE-Cy5)-conjugated anti-CD4 (GeneMay), phycoerythrin (PE)-conjugated anti-CD8
(Sigma-Aldrich), PE-conjugated anti-CD25 (GeneMay), FITC-conjugated anti-CD44
(GeneMay), FITC-conjugated anti-CD62L (GeneMay), FITC-conjugated anti-FOXP3
antibodies (eBioscience, CA, USA) and FITC-conjugated anti-IFN- antibodies
(eBioscience).
Cells to be immunostained were washed twice with 1x phosphate-buffered saline
(PBS) and suspended at a concentration of 1x106 cells/ml, followed by incubation in
the dark for 35 minutes at 4°C with appropriate volumes of two or three combinations
of fluorochrome-conjugated monoclonal antibodies (mAbs)
1x PBS.
For intracellular FOXP3 and IFN- staining, after cell surface molecules were stained
and washed with 1x PBS, 1ml freshly prepared Fixation/Permeabilization (1:3) working
solution (eBioscience) was added to each samples, followed by twice washing with 2ml
1x Permeabilization Buffer (eBioscience). The samples were then incubated with 20 l
corresponding antibody in the dark for at least 30 minutes at 4°C. Thereafter, cells
were washed twice with 2ml 1x Permeabilization Buffer, and then resuspended in Flow
Cytometry Staining Buffer (eBioscience).
Samples were analyzed by triple-color flow cytometry on a flow cytometer
(FACSCalibur, BD, NJ, USA). Cells were analyzed using CellQuest software (BD, USA).
In all cases at least 10,000 of gating events were required.
Statistical Analysis.
Statistical analysis was performed with Graphpad Prism
version 4.0 (Graphpad Software, Inc., San Diego, CA). Analysis of intergroup
differences was performed using Student’s t-test. A P-value < 0.05 was considered
statistically significant. All values were presented as mean ± standard error of
the mean (S.E.M.).
Results
Lymphodepletion after chemotherapy and subsequent recovery.
4
ml
of
peripheral blood was collected from NSCLC patients immediately prior to chemotherapy
(day -1), day 3 to 5, and day 7 to 9 post chemotherapy. Summarized data (n=21) showed
that the mean absolute number of leukocytes was reduced significantly from (7.8±0.5)
x 1012 at day -1 to (4.7±0.4) x 1012 at day 3 to 5 (P<0.0001), and was increased slightly
to (6.4±0.3) x 1012 at day 7 to 9 (P=0.0013). The mean absolute number of lymphocytes
was also decreased markedly from (2.1±0.1) x 1012 at day -1 to (1.0±0.1) x 1012 at
day 3 to 5 (P<0.0001). However, its absolute number was recovered to (1.6±0.1) x
1012 at day 7 to 9 (P<0.0001). Representation of the proportion of lymphocytes among
total leukocytes in percentages showed that the percentage of lymphocytes recovered
significantly from 20.3±1.3% at day 3-5 to 25.2±1.2% at day 7 to 9 (P=0.0093), a
level approaching that at day -1 (27.1±1.2%) (P=0.2747). This indicates that at least
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Dynamic change of immune pattern induced by chemotherapy
certain lymphocyte subsets recovered at a greater speed than the total leukocyte
population in the peripheral blood of cancer patients after depletion.
The reduction of lymphocyte proportion from day 3 to 5 is primarily attributed
to CD4+ T cells. Freshly isolated PBMCs from all patient populations were labeled
with a series of cell surface mAb markers (CD3, CD19, CD4, CD8) and multicolor flow
cytometric analysis were performed to further characterize these cells. Individual
frequencies for each of the lymphocyte subpopulations in all the patients analyzed
were represented as scatter plots. No significant differences were found between the
percentages of CD19+ B lymphocytes among total lymphocytes (n=11) throughout the
time-points observed. On the other hand, the percentage of CD3+ T lymphocytes among
total lymphocytes (n=14) was reduced significantly from 53.6±2.3% at day -1 to
42.0±2.8% at day 3 to 5 (P=0.0035), and almost restored to its pre-chemotherapeutic
level (52.1±2.7%) at day 7 to 9 (P=0.6774). Further gating of the CD3+ T lymphocytes
for CD4 and CD8 expression was carried out. The percentage of CD4+ T lymphocytes among
CD3+ T lymphocytes (n=17) plunged from 62.9±2.4% at day -1 to 54.4±2.4% at day 3
to 5 (P=0.0155), and recovered to 66.8±1.3% at day 7 to 8 (P<0.0001), a level similar
to its frequency before chemotherapy (P=0.1596). Again, analysis of the percentages
of CD8+ lymphocytes among CD3+ T lymphocytes (n=22) revealed no significant
differences throughout the time-points before and after chemotherapy. These data
indicate that the CD4+ lymphocyte subset is responsible for the trend of reduction
at day 3 to 5 followed by recovery of cellular proportion at day 7 to 8 observed in
flow cytometric analysis of peripheral blood lymphocytes.
Tregs exhibited a trend opposite to that of other subsets after chemotherapy in
lung cancer patients.
CD4+CD25+ are constitutively expressed by naturally
occurring regulatory T cells (9). As shown in Fig. 1A, the population of CD4+CD25+
lymphocytes as a percentage of total CD4+ cells was significantly reduced from
11.4±0.3% before chemotherapy to 8.2±0.4% at day 3 to 5 (P<0.0001) after
chemotherapy. Its frequency subsequently recovered to 10.4±0.5% at day 7 to 9
(P<0.0009), again a level with no significant difference compared to that before
chemotherapy (P=0.1007). Interestingly, CD4+CD25- lymphocytes, generally considered
to consist mainly of effector T cells carrying out immune response (30), showed a
trend opposite to that of CD4+CD25+ lymphocytes. Its frequency increased
significantly (P<0.0001) from an initial 88.6±0.3% to 91.8±0.4% at day 3 to 5,
followed by a decrease to 89.6±0.5% at day 7 to 9 (P=0.0009), a level similar to
that at day -1 (P=0.1013) (Fig. 1B). Representative flow cytometric data in Fig. 1C
showed the frequencies of these populations in the above time-points before and after
chemotherapy.
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论文集（2007）
Frequency of CD4 lymphocytes expressing CD25 in PBMCs of cancer patients (n=15) before
and after chemotherapy.
The CD4+ T population expressing CD25 (IL-2) are not entirely restricted to Tregs
and may also include activated T effector cells. Therefore we performed further gating
on the CD4+CD25+ populations for the expression of intracellular FOXP3 transcriptor
because FOXP3 is highly restricted to Tregs (10, 31) and regarded as the most reliable
marker for Tregs. Before chemotherapy (day -1), the CD4+CD25+FOXP3+ cells represented
6.7±0.4% of the total CD4+ T cell population in lung cancer patients, which was
significantly higher (P<0.0001) than that in healthy donors (n=10, 3.7±0.5%), as
shown in Fig. 2A, consisting with other reports that the level of Tregs was
significantly higher in cancer patients (18-20). However, representative flow
cytometric data showed that the population of CD4+CD25+FOXP3+ T cells was reduced
significantly at day 3 to 5 after chemotherapy but experienced a marked recovery at
day 7 to 9 (Fig. 2C). In Fig. 2A summarized scatter plot data of the 15 cancer patients
analyzed showed that the prevalence of this subset (Tregs) among CD4+ T cells was
reduced significantly (P<0.0001) from 6.7±0.4% at day -1 to 4.2±0.3% at day 3 to
5, a frequency comparable to that of healthy controls (P=0.3504), and restored to
a level similar to that before chemotherapy (P=0.4422) at day 7 to 9 (6.2±0.5%).
The frequencies of CD4+CD25+FOXP3- T cells did not show a significant increase or
decrease before and after chemotherapy (Fig. 2B).
This significant reduction of Tregs at day 3 to 5 and its subsequent recovery,
a stark contrast to that of CD25- and FOXP3- T cells, together with the marked decrease
of CD4+ T cells in comparison to other major lymphocyte subsets discussed above,
suggest that chemotherapy selectively depleted Tregs at day 3 to 5, and this
subpopulation restored to its original frequency at day 7 to 9.
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Dynamic change of immune pattern induced by chemotherapy
Fig. 2 Frequency of Tregs in PBMCs of cancer patients (n=15) before and after chemotherapy.
Summarised data and representative flow cytometric analysis of CD4+CD25+FOXP3+
lymphocytes (gated among CD4+ population) (A and B) displayed a trend showing Tregs
reducing significantly at d3~5 followed by subsequent recovery at d7~9. Comparison of
frequency of Tregs prior to chemotherapy and that in healthy donors (A) showed that this
population was significantly higher in lung cancer patients than in controls. The level
of CD4+CD25+FOXP3- population did not show any significant trend throughout the days
observed. P1, P2, P3, P4 and P5 are randomly selected patients in this analysis.
CD44highCD62Llow effector memory T cells underwent substantial expansion after
chemotherapy. CD44highCD62Llow T cells are widely regarded as effector memory T cells
(TEM) (32). To investigate the change in the prevalence of CD4+CD62Llow lymphocytes
after chemotherapy, we stained the PBMCs of cancer patients with anti-CD62L
immunofluorescent markers. Summarized data shown in Fig. 3A showed that the
CD4+CD62Llow population increased significantly from 3.0±0.4% at day -1 to 4.8±0.6%
at day 3 to 5 after chemotherapy (P=0.0392), but underwent a marked reduction to
2.6±0.5% at day 7 to 9 (P=0.0170). The frequency of CD62Llow cells among CD8+ T cells
also displayed a similar trend, increasing from 1.9±0.5% to 4.6±0.7% at day 3 to
5 (P=0.0121), and reducing to 2.6±0.5% at day 7 to 9 (P=0.0492) (Fig. 3B).
Further, flow cytometric analysis of surface expression of CD44 among total CD4+
T cells in cancer patients before and after chemotherapy showed a marked expansion
of this population from 11.9±2.2% at day -1 to 30.5±3.1% at day 3 to 5 (P<0.0001).
The frequency of this CD4+CD44high population maintains at this relatively high level,
and only decreased slightly to 27.5±3.0% at day 7 to 9 (P=0.4991) (Fig. 3C). Analysis
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of CD44 expression among total CD8+ cells showed a similar trend. As shown in Fig.
high
3E, the percentage of CD8+CD44
cells increased significantly (P=0.0153) from
10.4±0.8% before chemotherapy to 15.0±1.5% at day 3 to 5, and remains high through
day 7 to 9 (15.4±0.8%) (P=0.8195). Representative flow cytometric data in Fig. 3D
and Fig. 3F showed the frequencies of the CD4+CD44high and CD8+CD44high populations in
these time-points before and after chemotherapy.
high
low
Fig. 3 Substantial increase of CD44 CD62L
lymphocytes. Summarised data (n=10) and
low
representative flow cytometry data of both CD4+CD62L
cells (A and B) and CD8+CD62L
low
cells (C and D) showed a significant increase at d3~5, followed by a fall in frequency
at d7~9 (n=7). Analysis of CD44 expression among CD4+ (E) and CD8+ lymphocytes (F) showed
that these memory cells expanded substantially and markedly after chemotherapy at d3~5,
and remained at a high level through day 7 to 9. P1, P2, P3, P4 and P5 are randomly selected
patients in this analysis.
These data demonstrated that chemotherapy significantly expanded the CD62Llow
populations both in CD4 and CD8 T cells, but this subpopulation decreased sharply
at day 7 to 9. The CD44+ subsets in total CD4 and CD8 T cells also increased markedly
after chemotherapy at day 3 to 5, and these populations maintained a high level even
through day 7 to 9. Taken together, these data showed that the level of CD44highCD62Llow
T cells is at its maximum at day 3 to 5 after chemotherapy.
Intracellular production of IFN- showed continuous increase. IFN-
production by CD8+ cytotoxic T cells is one of the best described cytotoxic-mediated
immune responses. To assess cytolytic function, we performed intracellular flow
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Dynamic change of immune pattern induced by chemotherapy
cytometric analysis for IFN- in the PBMCs of lung cancer patients dynamically before
and after chemotherapy. As shown in representative flow cytometric data in Fig. 4,
the level of IFN- before administration of chemotherapeutic drugs in cancer patients
is similar to that of healthy controls, which mostly remains at a basal level of
0.05%~0.08% (Fig. 4). However, a significant up-regulation of IFN- intracellular
levels was detected in CD8+ T lymphocyte subpopulations. Intracellular IFN-
production exhibited a continuous and marked increase after chemotherapy, from
0.08±0.02% at day -1 to 0.56±0.15% (P=0.0153) at day 3 to 5 and up to 1.05±0.34%
at day 7 to 9 (P=0.2257) (Fig. 4A).
Fig. 4
Intracellular
cytokine
production of IFN- by CD8+ T
cells in peripheral blood of
lung cancer patients before and
after chemotherapy. Shown here
are summarised data (A) and
representative dot-plots (B) of
IFN- cytokine staining at day
-1, day 3~5, and day 7~9. The
level of IFN- production in
cancer patients prior to
chemotherapy is the same as that
in healthy donors (A).
Discussion
We have addressed for the first time, the dynamic change of frequencies and
intracellular IFN- production of various lymphocyte subpopulations after
chemotherapy in cancer patients. The present study contains the first evidence of
the time-phases at which the preferential depletion of Tregs induced by chemotherapy
and the subsequent recovery of this lymphocyte subset in cancer patients occur. It
also demonstrates the substantial expansion of CD44high CD62Llow-expressing lymphocyte
subpopulations, the latter’s frequency plunging back to its initial level before
chemotherapy, as well as the marked increase in IFN-, the time-phases of which
largely coincide with the reduction of Tregs.
In vivo immune condition after treatment with chemotherapeutic drugs had been
believed to be unfavorable for anti-tumor immunity induction due to their
lymphodepletive effects. On the other hand, tumor immunotolerance and escape from
immunosurveillance have long been a major hindrance for cancer immunotherapies, such
as the use of cancer vaccines and adoptive cell transfer therapy (33). The recent
analysis from Rosenberg et al reporting a regrettably low objective response rate
of 2.6% in 440 patients who underwent cancer vaccine trials in the National Cancer
Institute between 1995 and 2004 (34), has further demonstrated the need to develop
a better therapeutic approach.
The combination of chemotherapy (35, 36) or total-body irradiation (TBI) (37,
38) with cancer immunotherapies has, nevertheless, yielded relatively effective
anti-tumor immunity both in animal models (1, 2) and cancer patients (4-6), and has
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become a promising therapeutic approach for malignant neoplasms, compared with the
sole use of either treatment. It must be pointed out, however, that many of these
studies have only showed a marked expansion of tumor-antigen specific T lymphocyte
population, but with modest tumor regression or improvement of patient survival rate.
This illustrates the importance of clarifying the effect of chemotherapy on the immune
condition in cancer patients, that is, the mechanism by which chemotherapy favors
anti-tumor immunity induction, so that a better strategy of how to combine
chemotherapy with immunotherapies can be devised.
Such mechanism has thus far remained poorly elucidated. In cancer patients, only
Beyer et al in their study of CLL patients treated with fludarabine (39) and
Ghiringhelli et al in end stage cancer patients who underwent metronomic
cyclophosphamide regimen (40), has carried out analysis on the effect on chemotherapy
on Tregs frequency and functions. The answer to the question of how the frequencies
of this lymphocyte subset and that of the CD44highCD62Llow effector memory T cell subset,
as well as capacity of IFN- production, vary dynamically after chemotherapy has yet
to be provided, particularly after how many days does the recovery of Tregs take place
after its selective depletion and to what extent does the effector memory T cell subset
expands, such that a post-chemotherapeutic time phase at which the in vivo immune
environment is most favorable for anti-tumor immunity induction can be pinpointed.
The tumor immunity-suppressive function of Tregs in cancer patients have been
a heated area of research in recent years. This subset suppresses the activity of
a wide range of self/tumor antigen-reactive lymphocytic subpopulations (41); and this
study has demonstrated its increased frequencies in cancer patients, as shown also
by other colleagues (18-20). Attempts to selectively remove Tregs by drugs such as
humanized anti-Tac (anti-CD25) have limited applications due to their uncertain in
vivo effects (8). Hence non-specific lymphodepletion by immunosuppressive
chemotherapy and irradiation remains the only practical approach to eradicate Tregs
from cancer patients for the purpose of enhancing anti-tumor immunity (8).
Surprisingly, such lymphodepletive conditioning has been demonstrated to
selectively down-regulate Tregs frequencies and tumor immunity suppressive functions
(23, 39). More importantly, the change of Treg frequencies induced by chemotherapy
varies with time, and this subset can reconstitute to its original proportion after
a certain time-span post chemotherapy (42) and thus resumes its ability to suppress
tumor-specific CD4+CD25- T cells (43). Hence such down-regulation of Tregs induced
by chemotherapy is not persistent, and the delayed application of immunotherapy after
lymphodepletion can significantly compromise therapeutic outcome (44). Therefore
selecting a time point at which Tregs are reduced to a minimum after
chemotherapy-induced lymphopenia will be the key to therapeutic success of subsequent
cancer immunotherapies.
Our dynamic phenotyping of the PBMCs of cancer patients before and after
chemotherapy has led to the discovery of such time point – which is day 3 to day
5 after chemotherapy, a common trend observable in the majority of the individuals
of a sufficient sample size. The overall trend demonstrated that Tregs are reduced
to a greater extent compared to CD4+CD25- T cells, the latter even showed an increasing
trend in some patients. This time-point, we speculate, would be the optimum window
during which to carry out cancer immunotherapies such as tumor antigen vaccination
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Dynamic change of immune pattern induced by chemotherapy
and adoptive cell transfer therapy, at which maximal tumor regression can be achieved.
More importantly, our findings provide a valuable way of thought – induction of
active anti-tumor immunity should be attempted when Tregs are reduced but not after
this population recovered, the precise time-points of which should be analyzed with
similar approaches when the clinical trials are being carried out.
Lymphopenia-induced homeostatic proliferation, another active area of
investigation, enhances T-cell mediated tumor rejection by uncoupling tumor-specific
T cell anergy (45). During this process, CD44low naïve T cells interact with self/tumor
peptide MHC molecules (37) and convert to the CD44high memory phenotype (27).
Upregulation of this activation marker promote migration of lymphocytes through
extracellular matrix to tumor sites. Previous study has shown that cyclophosphamide,
another major chemotherapeutic agent, induces type I interferon and augments the
number of CD44high T lymphocytes in mice (46). Ma .et al, on the other hand, discovered
high
low
a significant increase in the percentage of CD44 CD62L effector memory T cells,
destined for recirculation from blood to tissue, following vaccination in TBI-treated
reconstituted lymphodepleted hosts, and these effector T cells demonstrated
increased tumor-specific immunity than those from normal hosts (38).
Our findings which show a trend of substantial and continuous increase of the
TEM subpopulations after chemotherapy-induced lymphodepletion, and a peaking of the
CD44highCD62Llow population at day 3 to 5, suggest this subset may derive from the skewing
of the T cell repertoire by homeostatic proliferation of naïve T cells into self/tumor
antigen-specific memory T cells, and serves to reconstitute the host’s immune system,
which may play a significant role in the active anti-tumor immune response and tumor
rejection induced by subsequent tumor antigens vaccination, and that such
proliferation activity may be most robust at day 3 to 5.
All in all, our study provides valuable data applicable in most clinical settings
of the suitable time-point for the use of immunotherapies after chemotherapy. More
importantly, we stress the importance of targeting the optimum opportunity by the
use of methods such as dynamic phenotyping of the subjects’ PBMCs to extract
information to be integrated into future therapeutic designs, for the use of such
combined therapies to achieve the best outcome.
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复旦大学
论文集（2007）
Acknowledgements
We would like to express our gratitude to the Chun-Tsung Undergraduate Research
Endowment established by Professor Lee Tsung-Dao for their grant and support to our
project.
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