Download Immunomodulatory effects of low dose chemotherapy and

IJC
International Journal of Cancer
Immunomodulatory effects of low dose chemotherapy and
perspectives of its combination with immunotherapy
Mariana S. Nars1 and Ramon Kaneno1,2
2
Department of Microbiology and Immunology, Institute of Biosciences, UNESP–Univ Estadual Paulista, Botucatu, São Paulo, Brazil
Department of Pathology, School of Medicine, UNESP–Univ Estadual Paulista, Botucatu, São Paulo, Brazil
Given that cancer is one of the main causes of death worldwide, many efforts have been directed toward discovering new
treatments and approaches to cure or control this group of diseases. Chemotherapy is the main treatment for cancer;
however, a conventional schedule based on maximum tolerated dose (MTD) shows several side effects and frequently allows
the development of drug resistance. On the other side, low dose chemotherapy involves antiangiogenic and
immunomodulatory processes that help host to fight against tumor cells, with lower grade of side effects. In this review, we
present evidence that metronomic chemotherapy, based on the frequent administration of low or intermediate doses of
chemotherapeutics, can be better than or as efficient as MTD. Finally, we present some data indicating that noncytotoxic
concentrations of antineoplastic agents are able to both up-regulate the immune system and increase the susceptibility of
tumor cells to cytotoxic T lymphocytes. Taken together, data from the literature provides us with sufficient evidence that low
concentrations of selected chemotherapeutic agents, rather than conventional high doses, should be evaluated in combination
with immunotherapy.
According to the most recent epidemiological surveys of the
World Health Organization, cancer is the third main cause of
death worldwide, just after cardiovascular diseases and infectious and parasitic diseases, being the second ranked disease
in most western countries.1 Data from the WHO and the
International Agency for Research on Cancer indicates that
12.7 million new cases were reported in 2008 and 7.6 million
deaths were attributed to this disease.2
The main types of human tumors comprise breast, lung,
prostate and colon cancer and the conventional treatment for
cancer includes surgery, chemotherapy and radiotherapy,
which can be used alone or in combination with each other.
Chemotherapy is the main modality of treatment being
administrated both in inoperable cases and as adjuvant treatment following surgery. In fact, more than half of all people
diagnosed with cancer receive chemotherapy as a neoadjuvant or adjuvant modality for preoperative or postoperative
treatment, respectively.3 In this review, we report the effects
of different antineoplastic chemotherapeutic agents on the
immune system, emphasizing the immunomodulatory effects
of moderate and low concentrations of drugs.
Key words: chemotherapy, metronomic, dendritic cells,
immunotherapy, Treg, vaccine
Abbreviations: BSA: body surface area-dosing; CEP: circulating
endothelial precursor cells; CTL: cytolytic T cells; CTX:
cyclophosphamide; DCs: dendritic cells; DOX: doxorubicin; LDM:
low dose chemotherapy; MDSC: myeloid-derived suppressor cells;
MTD: maximum tolerable dose; MTX: methotrexate; PTX: paclitaxel;
Treg: regulatory T lymphocytes
Grant sponsor: FAPESP–Fundação de Amparo a Pesquisa do
Estado de São Paulo; Grant numbers: 2009/18331-8 (regular
research grant) and 2009/18434-1 (scholarship)
Conflict of interest: Authors declare that there is no conflict of interest.
DOI: 10.1002/ijc.27801
Revised 29 Jun 2012; Accepted 16 Aug 2012; Online 28 Aug 2012
Correspondence to: Ramon Kaneno, Departamento de
Microbiologia e Imunologia, Instituto de Biociências de Botucatu,
UNESP. Cx Postal 510, Distrito de Rubião Jr. s/n, 18618-970,
Botucatu, São Paulo, Brazil, Tel.: [551438800432], Fax:
þ[55-14-38153744], E-mail: [email protected]
Conventional Chemotherapy
C 2012 UICC
Int. J. Cancer: 132, 2471–2478 (2013) V
Conventional chemotherapy is based on the maximum tolerated
dose (MTD), which corresponds to the highest dose of a drug
that presents tolerable side effects. This strategy has been shown
to be effective in curing or controlling the disease in a great
number of patients but it is associated with significant shortand long-term toxicity and side effects, including myelosuppression, neutropenia, thrombocytopenia, increased risk of infection
and bleeding, gastrointestinal dysfunctions, arthralgia, liver toxicity and damage to the cardiac and nervous system.3–5
One of the main features of antineoplastic chemotherapeutic agents is their cytotoxic activity on highly proliferative
cells, implying that its effect on proliferating hematopoietic
and immune cells leads to the depression of both natural and
adaptive defense mechanisms. In fact, the myelosuppressive
effect of several drugs is well described.6–10 As recently
reviewed by Shurin et al.10 the main immunosuppressive
effect of high dose chemotherapy is the depletion of T lymphocytes. Both CD4þ and CD8þ are affected by cytotoxic
Mini Review
1
Mini Review
2472
drugs but the normal levels of peripheral CD4þ cells are
more slowly restored than CD8þ.11,12 In addition, many
commonly used agents cause dysfunction of natural killer
(NK)13 and cdT cells,14 impairing the antitumor immune
surveillance. Although these cells are involved in the innate
immunity, the modulatory effect of paclitaxel (PTX), for
instance, seems to be dependent on the fail in their activation
by IL-2.15
Dendritic cells are also sensitive to high dose chemotherapy and the effects of several cytotoxic drugs have been
reported. In fact, such drugs can interfere on antigen presentation processes,16 reduce cell mobilization17 and downregulate the expression of cell surface markers such as CD80 and
CD86.18 Such effects on the immunocompetent cells may
enable tumor escape, thus allowing the proliferation of
chemo-resistant variants.19,20
To avoid extreme side effects and immunosuppression,
administration of cytotoxic drugs are followed by 3–4-week
intervals to allow hematopoiesis and restoration of peripheral
leukocyte count. However, the low blood levels or absence of
medication during these resting intervals can also allow the
growth of drug-resistant tumor cell clones. Then, despite an
initially impressive tumor regression or even remission following chemotherapy, regrowth or recurrence is quite common in
metastatic cancer.21 As a consequence, it is estimated that 45%
of cancer patients are unable to control the disease.22
MTD is calculated by the body surface area-dosing (BSA)
method, a calculation that can lead to an unpredictable variation in the therapeutic effect, due to either overdosing or
underdosing. Overdosing is easily detected due to the
increased toxicity, whereas underdosing is more difficult to
recognize and may occur in 30% or more of patients receiving standard regimen. Therefore, those underdosed patients
are at risk of a significantly reduced anticancer effect of the
medication.23 For instance, a survival reduction of almost
20% was observed in breast cancer patients treated with cyclophosphamide (CTX), doxorubicin and 5-fluorouracil at
adjuvant BSA-based concentrations.24,25 The authors considered the possibility that a large proportion of these patients
had been underdosed, which may have facilitated the development of drug-resistance.
Several mechanisms can be associated with the development of drug resistance including the expression of ATPbinding cassette family transporters, that can exclude cytotoxic drugs from target cells26,27; development of resistance to
apoptosis related to Bcl-228,29 and Notch receptor expression30,31 as well as immunosuppression32–34 that can allow
the development of ‘‘dormant tumor stem cells.’’ Another important phenomenon for developing drug resistance is the
epigenetic changes in tumor cells. For instance, resistance of
colorectal tumor cells to 5-fluorouracil (5-FU) is hypothesized to be acquired by DNA hypermethylation. In fact, treatment of 5-FU-resistant colon cancer cells (HCT-8) with low
dose of 5-aza-deoxycytidine, prevents DNA hypermethylation
and restores their sensitivity to 5-FU.35 This effect was also
Immunomodulation by low dose chemotherapy
demonstrated in vivo by treating nude mice bearing 5-FUresistant tumor cells. Treatment with low doses of AZA overcame the resistance to 5-FU.36 Therefore, there is a great interest in minimizing toxicity while increasing drug efficacy,37
suggesting the necessity for different approaches.
Low Dose Metronomic Chemotherapy
One of the alternative approaches is low-dose metronomic
(LDM) chemotherapy, which uses much lower doses of chemotherapeutics and may decrease the disadvantages of conventional cytotoxic chemotherapy.19,20 The difference
between this approach and simple underdosing is that the
former involves more frequent administration38–40 to maintain the serum levels of antineoplastic activity and to avoid
myelosuppression and other side effects. It has attracted
attention given that it may reduce the risk of developing
drug resistance while reducing the toxic effects.41
One of the main antitumor mechanisms of LDM is the
antiangiogenic activity, since endothelial precursor cells are
more sensitive to cytotoxic drugs than tumor cells themselves.42,43 Such effect was shown to be enhanced in vivo by
combination with antiangiogenic agents. In fact, it was
observed in animal models that the association of antineoplastic drugs with antiangiogenic agents such as anti-VEGF, antiVEGFR, b-cyclodextrin tetradecasulfate, tetrahydrocortisol and
endoglin significantly increased the survival of tumor-bearing
animals.44–47 The absence of side effects and a visible decrease
in tumor size were associated with the treatments.44,48,49 MTD
and LDM chemotherapy have opposite effects on the mobilization and viability of circulating endothelial precursor (CEP)
cells, with LDM being more effective than MTD at destroying
these cells involved in tumor angiogenesis.50
In vitro antiangiogenic activity of LDM docetaxel has also
been shown, while its in vivo effectiveness was demonstrated
in a murine gastric tumor model. It was observed that it
affected microvessel growth without exerting a toxic effect on
nude mice.51
Interestingly, looking back at the history of pediatric cancer therapy, several successful approaches could be classified
as metronomic chemotherapy but instead are known as
‘‘maintenance therapy.’’37 For instance, pediatric oncologists
have shown that daily mercaptopurine and weekly methotrexate administrations constitute a successful combination
against leukemia.20 The main knowledge on LDM is based
on the studies with CTX and PTX, but the feasibility of using
other medications alone or in combination has also been
investigated.
Cyclophosphamide
CTX is an alkylating agent that is converted into its active
metabolite, derivative phosphoramide mustard, in the liver.
This metabolite inhibits DNA replication by forming interstand and intrastrand DNA crosslinking. Currently, it is the
best-known clinical metronomic drug52 and it was observed
that LDM CTX increases the frequency of apoptotic CEPs,
C 2012 UICC
Int. J. Cancer: 132, 2471–2478 (2013) V
while high-dose CTX mobilizes viable ones giving a significant contribution to the growth of experimental lymphoma.53
Examples of the clinical use of LDM CTX include the combination of weekly vincristine and low-dose daily CTX that was
shown to be effective against neuroblastoma.54 Combination
of LDM CTX with celecoxib in heavily pretreated patients
contributed to an impressive duration of stable disease with
no toxicity,45 whereas this combination is also effective in
treating refractory high-grade lymphoma.55 The combination
of CTX plus weekly vinblastine and rofecoxib is effective for
a subset of Hodgkin’s disease patients.56 CTX has also been
used with low-dose methotrexate in the treatment of early
breast cancer and the inhibition of angiogenesis is caused by
a decrease in vascular endothelial growth factor.57
Effects of CTX on the immune response and combination
with immunotherapy
Regulatory T cells and myeloid-derived suppressor cells
(MDSC) are in higher number in the tumor microenvironment and can facilitate tumor immune escape, since they
affect the functioning of effective CTLs and NK cells.58,59 Tumor-bearing mice show an increased prevalence of Tregs not
only at the tumor site but also in the peripheral blood and
lymph nodes when compared with normal controls.60–62 Otherwise, selective depletion of these cells may be accomplished
following low-dose CTX administration. The administration
of this drug at 45-day intervals block the renewal of Tregs in
mice with multiple myeloma and enable the restoration of an
efficient immune response against the tumor cells, thereby
leading to prolonged survival and prevention of disease recurrence. Then, low-dose CTX induces beneficial immunomodulatory effects in the immunotherapy context,61–63 but
more frequent administrations at 7- or 21-day intervals did
not improve the therapeutic effect, since the mice developed
multiple myeloma at a higher incidence.51
Intraperitoneal injection of CTX significantly suppresses
bilateral growth of murine colon cancer cells (CT-26). Association of this drug with doxorubicin decreased the tumor
growth inducing the remotely implanted cells, indicating the
development of systemic tumor-specific T cell response.64 In
fact, such combination increased the frequency of IFN-c-producing cells and decreased that the expression of FoxP3 and
TGF-b within the remote tumor site.
Distinctive time-schedule administration of low-dose CTX
is beneficial for breaking immune tolerance while its modulatory effect on DC-based immunotherapy leads to increased
survival of tumor-bearing mice, which can be further
improved by elimination of Tregs.65 These results suggest
that the DC-based antitumor treatment of patients might be
improved by simultaneous depletion of Tregs using a low
dose of CTX. Moreover, this combination significantly
improves the survival of tumor-bearing mice, when compared
with DC vaccination or CTX alone. Despite the well-documented effects of CTX on Treg elimination, these cells are
rapidly reconstituted in both secondary lymphoid tissues and
C 2012 UICC
Int. J. Cancer: 132, 2471–2478 (2013) V
2473
at the tumor site, which could discourage new approaches in
this field. Further attempts to prolong Treg depletion have
yielded no additional benefit.66
It is generally believed that the antitumor immunity
induced by the combination of chemotherapy and immunotherapy depends on the homeostatic role of immunotherapy
acting during the recovery phase that follows the chemotherapy mediated myelolymphodepletion.67 The ability of this
drug to reverse the systemic and in situ DC paralysis in
tumor-bearing hosts and its ability to mobilize early proliferating DC progenitors are other crucial mechanisms for enhancing antitumor immunity.66 In this aspect, it was observed that
CTX administration restores myelopoiesis, leading to reversion
of systemic and in situ tumor-induced DC paralysis. Moreover,
CTX mobilized proliferating early DC progenitors to the tumor site where they yielded new DCs.
These several findings are relevant for understanding the
multifaceted mechanisms behind the beneficial effects of CTX
on augmenting antitumor immunity. In other words,
although this drug shows myelosuppressive activity and eliminates the tumor-infiltrating DCs, it is able to mobilize early
DC precursors. These precursor cells proliferate rapidly on
the periphery and increase the presence of new APCs that
can prime de novo T cell responses.66
Thus, CTX has a direct impact on DC-T cell interactions
in a tumor-bearing host that ultimately lead to a better priming of tumor-specific T cells. These observations support
the notion that mobilization of early DC precursors to sites
of ongoing immune response and their differentiation into
mature DCs can lead to optimal T cell priming.68,69 Therefore, CTX can influence the DC homeostasis both in situ and
systemically through its unique ability to mobilize DC
precursors.66
Another important finding on the immune effects of CTX
was that a single administration of low-dose CTX (50 mg/kg)
in tumor-bearing mice prior to immunization with DC
increases the frequency of IFN-c secreting antitumor CTLs.61
This is in agreement with the fact that different chemotherapeutic agents at very low concentrations increase the ability
of DCs to induce T cell proliferation.70
Considering that the use of monoclonal antibodies is the
more prominent modality of immunotherapy, Garcia et al.71
have reported remarkable findings in a phase II study of lowdose CTX and bevacizumab against recurrent, refractory
ovarian cancer. They have shown that the administration of
CTX increased the probability of progression free survival of
bevacizumab-treated patients. Combination was not free of
toxicity and indeed, lymphopenia was one of most common
side effects showed by patients.
Several groups have made efforts to develop safe and
effective antitumor vaccines using cytokine genes inserted
into viral vectors. In this direction, Malvicini et al.72 observed
in a murine model of colorectal carcinoma that the combination of CTX with an IL-12-encoding adenovirus vector induces complete regression in more than 50% of tumor-bearing
Mini Review
Nars and Kaneno
2474
Mini Review
mice. Such treatment was able to decrease the number of peripheral and spleen Treg, at the same time that it increased
the numbers of activated DC and IFN-c-secreting CD4þ
lymphocytes.
The beneficial effects of CTX in augmenting antitumor
immunity can be due to the combination of its ability to
eliminate Tregs, reset the DC homeostasis in tumor-bearing
host and restore the homeostasis-driven expansion of preexisting tumor-specific effector T cells.60,73,74
Paclitaxel
PTX is a mitotic inhibitor that causes cell cycle arrest by stabilizing tubulin in microtubules. It is another useful agent for
LDM therapy given its broad spectrum antitumor activity
and ability to in vitro inhibit endothelial cell functions that
are fundamental for angiogenesis. In fact, comparing MTD
and LDM therapies with PTX, it can be observed that the latter presents stronger antiangiogenic and antilymphangiogenic
activities than the former in breast cancer metastases. Since
breast cancers spread predominantly through the lymphatic
system,75 the antilymphangiogenic activity of LDM PTX can
be considered a fundamental feature of this schedule. This
drug overcomes the angiogenic role of VEGF-A and reduces
the rate of prostate cancer growth in rats by 24% and the
vascularized area by 35%.41 However, the benefits of LDM
and antiangiogenic therapies are generally limited by
acquired resistance76; since, these regimens might lead to
selection of hypoxia-resistant tumor cell populations as a
mechanism of tumor escape.77 In addition, patients under
LDM PTX show less pronounced side effects such as body
weight loss and leukopenia than those treated with MTD.
This reduced toxicity makes it possible to use LDM as an adjuvant therapy for prolonged periods against advanced metastatic breast cancer.78
Immunomodulatory effects of PTX and combination with
immunotherapy
In a murine lung carcinoma model, the combination of lowdose PTX followed by intratumoral injection of DC vaccine
is much more efficient for the treatment than either DC vaccine or PTX alone. The combination was shown to inhibit
tumor growth, induce tumor infiltration by CD4þ and
CD8þ T cell and induce a tumor-specific immune response
in regional lymph nodes. The chosen concentration of PTX
showed no toxicity to bone marrow cells and, in addition,
was able to stimulate DC maturation and function and prevent the inhibitory activity of tumor cells on DC maturation
and motility.67 These observations supported the design of
intratumoral administration of DCs after pretreatment of tumor-bearing mice with LDM PTX.79 Such combination
inhibits the s.c. development of murine lung cancer, increasing the infiltration of CD4þ and CD8þ cells. Increased survival was also correlated with release of IFN-c by draining
lymph nodes lymphocytes and local production of MCP-1
and IP-10. Similar results were observed in the tumor pro-
Immunomodulation by low dose chemotherapy
gression of a breast cancer model, which growth was delayed
in contrast with the poor activity of drug alone.80
Immunotherapy based on the administration of cytokines
is one of the goals of several groups. It was observed, for
instance, that the incubation of tumor cells with granulocytemacrophage colony-stimulating factor (GM-CSF) induces cell
surface modification.81
Low-dose PTX combined with the GM-CSF-surface-modified tumor-cell vaccine effectively enhances tumor regression
when the vaccine is administered after PTX. This drug
mainly enhances the antitumor immune response at the priming time, being the administration of the lower doses (4
and 20 mg/kg) prior to the vaccine superior to post-vaccine
treatment. However, the frequency of administrations
depends on the dose of the drug, since at higher doses (40
mg/kg) no significant differences were identified between the
administration sequences.81 The administration of the lowdose PTX followed by the vaccine induced a high degree of
CD8þ T cell infiltration in tumor tissue, suggesting the
induction of tumor-specific immune response. The combination treatment also induced a higher number of tumor-specific IFN-c CD8þ T cells than vaccine alone. Low-dose PTX
is sufficiently potent to induce phagocytosis of tumor antigens by DCs and to enhance their maturation and priming
of antitumor immune response.81
It is important to note that high PTX concentrations
induce DC apoptosis, whereas low concentrations not only
diminish apoptosis of these cells but also block the inhibitory
effect of the tumor on DC maturation. The effect of this type
of treatment may be enhanced by selecting the proper
sequence for various drug-vaccine combinations based on the
immunomodulatory mechanisms of each treatment.81 Moreover, MTD is more effective at inducing tumor apoptosis and
there is a significant difference between these two schedules
concerning the apoptosis index of tumor cells.82 However,
even though the destruction of tumor cells is the main goal
of the treatments, lack of apoptosis can play a favorable role
as will be discussed below.
Other chemotherapeutic agents
Effects of metronomic therapy with other common chemotherapeutics such as 5-FU, gencitabine and oxaliplatin have
also been evaluated. In contrast with CTX, gencitabine
showed no effect on Treg, but selectively reduces MDSC
(CD11bþ/Gr1þ cells)83,84 and enhances the antitumor activity of CD8þ T and NK cells.84
Correale’s group has analyzed the efficacy of these drugs
associated with monoclonal antibodies based immunotherapy
and reported that treatment of high-risk non-small cell lung
cancer patients with cisplatin plus etoposide promoted total
or partial remission of disease in 45.2% of the patients enrolled in a phase II trial. Similarly, to previously described
drugs, the activity of this metronomic schedule was attributed
to changes on tumor neoangiogenesis.85 Such effect was further reported to be enhanced by combination with the antiC 2012 UICC
Int. J. Cancer: 132, 2471–2478 (2013) V
VEGF humanized antibody bevacizumab.86 However, this
chemoimmunotherapy combination was not completely safe
and some patients presented side effects such as hematological changes, mucosal toxicity, alopecia, infections, thromboembolia and mood depression.86,87
Another chimeric antibody Cetuximab, directed to the receptor for EGF, was associated with irinotecan, leucovorin
and 5-FU for treating colon cancer patients. This combination was able to increase the expression of antigens by tumor
cells. Immunological effects included increased CTL activity,
probably due to the enhanced susceptibility of tumor cells to
endocytosis by DC.88
Immunomodulatory Effects of Ultra-Low
Concentrations of Chemotherapeutics
It was observed that ultra-low noncytotoxic concentrations of
selected antineoplastic agents are able to modulate the
immune system. Recent findings suggest that some chemotherapeutic drugs can improve the efficacy of immunotherapy
by different means. Therefore, many efforts have been
directed toward the development of immunotherapeutic
approaches to enable the immune system to respond specifically against cancer.20,89
Dendritic cells have different sensitivities to conventional
cytotoxic agents. In this aspect, it was reported that PTX and
other chemotherapeutic agents could up-regulate DC functions in vitro.79 Studies from our group showed that low
noncytotoxic concentrations of doxorubicin, methotrexate,
mitomycin C and PTX directly up-regulate the ability of DCs
to present antigens for Ag-specific T cells in vitro. These
drugs significantly up-regulate the expression of CD80,
CD86, CD40 and MHC class II molecules on DCs.90 In addition, we observed that chemotherapeutic agents up-regulate
APC function of DCs. In vitro treatment of DCs with doxorubicin, PTX or methotrexate increases their ability to present
ovalbumin (OVA) to OVA-specific CD8þ T cells.90 This
treatment also up-regulates the expression of the antigen
processing machinery (APM) components, including immunoproteasomes (LMP7 and LMP10) and chaperones such as
tapasin and calmodulin (delta).90
Furthermore, IL-12 might also play an important role in
this phenomenon as a signal 3 molecules91,92 and, indeed,
doxorubicin (DOX), methotrexate (MTX), PTX and vinblastine increase spontaneous IL-12 expression in treated DCs.90
In fact, in a murine model, the increased ability of DCs to
present antigens to Ag-specific T cells after treatment with
such drugs is abolished in DCs generated from IL-12 knockout mice. It indicates that up-regulation of this phenomenon
is IL-12-dependent and mediated by autocrine or paracrine
mechanisms. As like as human DC, murine cells are also up
regulated by doxorubicin methotrexate and PAC to express
the mature DC phenotype and increase their ability to stimulate the allogeneic response in the mixed lymphocyte reaction.70 Interestingly, higher concentrations of mitomycin C
(up to 6.0 lM) induce the generation of tolerogenic DCs,
C 2012 UICC
Int. J. Cancer: 132, 2471–2478 (2013) V
2475
which express low levels of CD80 and CD86 and display
weak activity in the MLR assay.18
Increased expression of CD40 molecules on DCs treated
with methotrexate and mitomycin C is in agreement with
their increased ability to stimulate T cell proliferation in the
MLR assay. However, this correlation was not seen for other
tested drugs, suggesting the importance of other mechanisms
involved in up-regulation of antigen-presenting function of
DCs by chemomodulation.70 Expression of CD40 on DCs is
essential for their interaction with T lymphocytes and development of efficient Th1 responses.93 Because expression of
CD40 on DCs and the CD40-mediated DC function are suppressed during tumor progression,94 its up-regulation by nontoxic chemotherapy should support the development of antitumor immunity in tumor-bearing hosts. In addition, CD40
ligation protects human and murine DCs from tumor-induced
apoptosis by inducing expression of antiapoptotic proteins
from the Bcl-2 family.95–97 Therefore, a combination of chemotherapy plus immunotherapy has been discussed and requires
more studies to support its widespread clinical use.
Development of spontaneous melanoma in ret transgenic
mice can be inhibited by their treatment with low dose
PTX.98 This effect was observed to be associated with lower
number of infiltrating MDSC in the tumor microenvironment. Michels et al.99 reported similar findings in C57BL/6
mice with PTX slightly decreasing the generation of MDSC
from bone marrow precursor cells. Such decrease was not
due to induction of apoptosis of these cells. It reinforces the
evidence that PTX is able to modulate the immune system.
We also investigated the effect of ultra-low PTX concentrations on colon cancer cells. First, the DNA microarray
showed that treatment of tumor cells induced transcriptional
changes in several genes associated with both tumor immunogenicity and cell growth. Analysis of APM components of
these cells showed that the treatment up regulated the expression of proteasomes, TAP, tapasin and calnexin. These results
suggested to us that the tumor immunogenicity could be
affected by such changes, which were confirmed by the
increased ability of treated tumor cells to induce the generation of tumor-specific cytolytic T lymphocytes. In addition,
PTX-treated tumor cells showed higher susceptibility to CTL
activity than the wild type cells.100 This finding was reinforced by other authors who have observed that the in vitro
treatment of murine colon cancer cells with low concentrations of PTX or doxorubicin increased their sensitivity to
specific anti-p53 CTL.80 Authors obtained similar results
treating human lung cancer cells (H332). Although no direct
effect of PTX on murine lymphocytes was observed, doxorubicin negatively affected cell toxicity80 and human peripheral
blood cells were sensitive to drug treatment. Such enhanced
sensitivity of tumor cells to CTL cytotoxicity was attributed
to the ability of PTX, doxorubicin and cisplatin to increase
the cell permeability to Granzyme B.80
The dying tumor cells can release danger signals (damageassociated molecular pattern, DAMP) that lead to the
Mini Review
Nars and Kaneno
Mini Review
2476
Immunomodulation by low dose chemotherapy
activation of DCs and may facilitate the engulfing and processing of tumor antigens101,102; however, it is usually supposed that massive tumor cell death or apoptosis is nonimmunogenic.
The
molecular
pathways
of
certain
‘‘immunogenic’’ forms of cancer cell death that cause DC
activation have recently been defined. In this aspect, it was
reported that DNA-damaging anthracyclines (idarubicin,
doxorubicin and mitoxantrone) can induce the translocation
of calreticulin to the surface of dying tumor cells, thus promoting phagocytosis by DCs. The surface exposure of calreticulin is an event that occurs only in immunogenic death,
making it an important mediator in the uptake of tumor cells
by DCs.103 Some danger signals, such as the high-mobility
group box 1 (HMGB1) alarmin protein, are secreted by dying
tumor cells, which interact with Toll-like receptor 4 (TLR4)
expressed by DCs.
In addition, many chemotherapeutic agents, including
CTX, induce production of uric acid caused by tumor cell
death, which may activate DCs, thereby promoting tumor
rejection.104 Moreover, the ability of vinblastine-treated
mature DCs to induce CD8þ T cell responses is higher,
when compared with untreated DCs.105
Direct effect of noncytotoxic concentrations of drugs on lymphocytes has not been reported except that doxorubicin can
affect murine CTL, whereas PTX, doxorubicin and cisplatin can
reduce the activity of human cytotoxic cells.80 Our findings on
this subject indicate that lymphocytes are resistant to noncytotoxic concentrations of 5-fluorouracil, 5-aza-deoxycytidine and
PTX. Differently of DC, lymphocyte functions are not modulated by such low concentrations of drugs (unpublished data),
making DC the main target for chemoimmunemodulation.
Conclusion and Perspectives
Low-dose chemotherapy, including the use of noncytotoxic
concentrations, metronomic schedule and noncontinuous
low-dose administration of antineoplastic agents is an alternative for cancer chemotherapy. These different schedules
minimize the side effects of conventional MTD and reduce
the rise of resistant variants of tumor cells, which are the
main disadvantages of classical treatment. The up-regulating
effects on DCs allow us to suggest that the use of low concentrations of selected chemotherapeutics, rather than high
doses, should be used in combination with immunotherapy
to overcome the immunosuppression induced by cancer.
References
1.
2.
3.
4.
5.
6.
7.
8.
9.
10.
World Health Organization. Global health risks:
mortality and burden of disease attributable to
selected major risks. Geneva: WHO Press, 2009.
70p. Available: http://www.who.int/healthinfo/
global_burden_disease/GlobalHealthRisks_
report_full.pdf (accessed on August 1, 2012).
International Agency for Research on Cancer.
Globocan 2008 Fast Stats. Available at: http://
globocan.iarc.fr/factsheets/populations/factsheet.
asp?uno=900#BOTH (accessed on August 1, 2012).
Andre T, de Gramont A. An overview of
adjuvant systemic chemotherapy for colon
cancer. Clin Colorectal Cancer 2004;4 Suppl 1:
S22–8.
McWhinney SR, Goldberg RM, McLeod HL.
Platinum neurotoxicity pharmacogenetics. Mol
Cancer Ther 2009;8:10–6.
Vento S, Cainelli F, Temesgen Z. Lung
infections after cancer chemotherapy. Lancet
Oncol 2008;9:982–92.
Rasmussen L, Arvin A. Chemotherapy-induced
immunosuppression. Environ Health Perspect
1982;43:21–5.
Wijayahadi N, Haron MR, Stanslas J, et al.
Changes in cellular immunity during
chemotherapy for primary breast cancer with
anthracycline regimens. J Chemother (Florence,
Italy) 2007;19:716–23.
Naiditch H, Shurin MR, Shurin GV. Targeting
myeloid regulatory cells in cancer by
chemotherapeutic agents. Immunol Res 2011;50:
276–85.
Kim SK, Demetri GD. Chemotherapy and
neutropenia. Hematol Oncol Clin North Am
1996;10:377–95.
Shurin MR, Naiditch H, Gutkin DW, et al.
Chemoimmunomodulation: immune regulation
by the antineoplastic chemotherapeutic agents.
Curr Med Chem 2012;19:1792–803.
11.
12.
13.
14.
15.
16.
17.
18.
Mackall CL, Fleisher TA, Brown MR, et al.
Distinctions between CD8þ and CD4þ T-cell
regenerative pathways result in prolonged T-cell
subset imbalance after intensive chemotherapy.
Blood 1997;89:3700–7.
Azuma E, Nagai M, Qi J, et al. CD4þ Tlymphocytopenia in long-term survivors
following intensive chemotherapy in childhood
cancers. Med Pediatr Oncol 1998;30:40–5.
Alanko S, Salmi TT, Pelliniemi TT. Recovery of
natural killer cells after chemotherapy for
childhood acute lymphoblastic leukemia and
solid tumors. Med Pediatr Oncol 1995;24:
373–8.
Zocchi MR, Poggi A. Role of gammadelta T
lymphocytes in tumor defense. Front Biosci
2004;9:2588–604.
Chuang LT, Lotzova E, Heath J, et al. Alteration
of lymphocyte microtubule assembly,
cytotoxicity, and activation by the anticancer
drug taxol. Cancer Res 1994;54:
1286–91.
Nakashima H, Tasaki A, Kubo M, et al. Effects
of docetaxel on antigen presentation-related
functions of human monocyte-derived dendritic
cells. Cancer Chemother Pharmacol 2005;55:
479–87.
Ferrari S, Rovati B, Porta C, et al. Lack of
dendritic cell mobilization into the peripheral
blood of cancer patients following standard- or
high-dose chemotherapy plus granulocytecolony stimulating factor. Cancer Immunol
Immunother 2003;52:359–66.
Jiga LP, Bauer TM, Chuang JJ, et al. Generation
of tolerogenic dendritic cells by treatment with
mitomycin C: inhibition of allogeneic T-cell
response is mediated by downregulation of
ICAM-1, CD80, and CD86. Transplantation
2004;77:1761–4.
19.
20.
21.
22.
23.
24.
25.
26.
27.
28.
Schlom J, Arlen PM, Gulley JL. Cancer vaccines:
moving beyond current paradigms. Clin Cancer
Res 2007;13:3776–82.
Nowak AK, Lake RA, Robinson BW. Combined
chemoimmunotherapy of solid tumours:
improving vaccines? Adv Drug Deliv Rev 2006;
58:975–90.
Hahnfeldt P, Folkman J, Hlatky L. Minimizing
long-term tumor burden: the logic for
metronomic chemotherapeutic dosing and its
antiangiogenic basis. J Theor Biol 2003;220:
545–54.
Pirozynski M. 100 years of lung cancer. Respir
Med 2006;100:2073–84.
Gurney H. How to calculate the dose of
chemotherapy. Br J Cancer 2002;86:1297–302.
Budman DR, Berry DA, Cirrincione CT, et al.
Dose and dose intensity as determinants of
outcome in the adjuvant treatment of breast
cancer. The Cancer and Leukemia Group B. J
Natl Cancer Inst 1998;90:1205–11.
Silber JH, Fridman M, DiPaola RS, et al. Firstcycle blood counts and subsequent neutropenia,
dose reduction, or delay in early-stage breast
cancer therapy. J Clin Oncol 1998;16:2392–400.
Baguley BC. Novel strategies for overcoming
multidrug resistance in cancer. BioDrugs 2002;
16:97–103.
Chen KG, Valencia JC, Lai B, et al.
Melanosomal sequestration of cytotoxic drugs
contributes to the intractability of malignant
melanomas. Proc Natl Acad Sci USA 2006;103:
9903–7.
Michaud WA, Nichols AC, Mroz EA, et al. Bcl2 blocks cisplatin-induced apoptosis and
predicts poor outcome following chemoradiation
treatment in advanced oropharyngeal squamous
cell carcinoma. Clin Cancer Res 2009;15:
1645–54.
C 2012 UICC
Int. J. Cancer: 132, 2471–2478 (2013) V
29.
30.
31.
32.
33.
34.
35.
36.
37.
38.
39.
40.
41.
42.
43.
44.
45.
46.
Pham CG, Bubici C, Zazzeroni F, et al.
Upregulation of Twist-1 by NF-kappaB blocks
cytotoxicity induced by chemotherapeutic drugs.
Mol Cell Biol 2007;27:3920–35.
Nefedova Y, Sullivan DM, Bolick SC, et al.
Inhibition of Notch signaling induces apoptosis
of myeloma cells and enhances sensitivity to
chemotherapy. Blood 2008;111:2220–9.
Li M, Chen F, Clifton N, et al. Combined
inhibition of Notch signaling and Bcl-2/Bcl-xL
results in synergistic antimyeloma effect. Mol
Cancer Ther 2010;9:3200–9.
Baguley BC, Finlay GJ, Ching LM. Resistance
mechanisms to topoisomerase poisons: the
application of cell culture methods. Oncol Res
1992;4:267–74.
Demicheli R, Retsky MW, Hrushesky WJ, et al.
Tumor dormancy and surgery-driven
interruption of dormancy in breast cancer:
learning from failures. Nat Clin Pract Oncol
2007;4:699–710.
Koebel CM, Vermi W, Swann JB, et al. Adaptive
immunity maintains occult cancer in an
equilibrium state. Nature 2007;450:903–7.
Humeniuk R, Mishra PJ, Bertino JR, et al.
Epigenetic reversal of acquired resistance to 5fluorouracil treatment. Mol Cancer Ther 2009;8:
1045–54.
Jabbour E, Issa JP, Garcia-Manero G, et al.
Evolution of decitabine development:
accomplishments, ongoing investigations, and
future strategies. Cancer 2008;112:2341–51.
Baruchel S, Stempak D. Low-dose metronomic
chemotherapy: myth or truth? Onkologie 2006;
29:305–7.
Bocci G, Nicolaou KC, Kerbel RS. Protracted
low-dose effects on human endothelial cell
proliferation and survival in vitro reveal a
selective antiangiogenic window for various
chemotherapeutic drugs. Cancer Res 2002;62:
6938–43.
Kamen BA. Metronomic therapy: it makes sense
and is patient friendly. J Pediatr Hematol Oncol
2005;27:571–2.
Miller KD, Sweeney CJ, Sledge GW, Jr.
Redefining the target: chemotherapeutics as
antiangiogenics. J Clin Oncol 2001;19:1195–206.
Lennernas B, Albertsson P, Damber JE, et al.
Antiangiogenic effect of metronomic paclitaxel
treatment in prostate cancer and non-tumor
tissue in the same animals: a quantitative study.
APMIS 2004;112:201–9.
Browder T, Butterfield CE, Kraling BM, et al.
Antiangiogenic scheduling of chemotherapy
improves efficacy against experimental drugresistant cancer. Cancer Res 2000;60:1878–86.
Klement G, Baruchel S, Rak J, et al. Continuous
low-dose therapy with vinblastine and VEGF
receptor-2 antibody induces sustained tumor
regression without overt toxicity. J Clin Invest
2000;105:R15–24.
Bello L, Carrabba G, Giussani C, et al. Low-dose
chemotherapy combined with an antiangiogenic
drug reduces human glioma growth in vivo.
Cancer Res 2001;61:7501–6.
Klement G, Huang P, Mayer B, et al.
Differences in therapeutic indexes of
combination metronomic chemotherapy and an
anti-VEGFR-2 antibody in multidrug-resistant
human breast cancer xenografts. Clin Cancer Res
2002;8:221–32.
Soffer SZ, Moore JT, Kim E, et al. Combination
antiangiogenic therapy: increased efficacy in a
47.
48.
49.
50.
51.
52.
53.
54.
55.
56.
57.
58.
59.
60.
61.
C 2012 UICC
Int. J. Cancer: 132, 2471–2478 (2013) V
murine model of Wilms tumor. J Pediatr Surg
2001;36:1177–81.
Teicher BA, Sotomayor EA, Huang ZD.
Antiangiogenic agents potentiate cytotoxic
cancer therapies against primary and metastatic
disease. Cancer Res 1992;52:6702–4.
Tan GH, Tian L, Wei YQ, et al. Combination of
low-dose cisplatin and recombinant xenogeneic
endoglin as a vaccine induces synergistic
antitumor activities. Int J Cancer 2004;112:
701–6.
Hou JM, Liu JY, Yang L, et al. Combination of
low-dose gemcitabine and recombinant quail
vascular endothelial growth factor receptor-2 as
a vaccine induces synergistic antitumor
activities. Oncology 2005;69:81–7.
Untergasser G, Koeck R, Wolf D, et al. CD34þ/
CD133- circulating endothelial precursor cells
(CEP): characterization, senescence and in vivo
application. Exp Gerontol 2006;41:600–8.
Wu H, Xin Y, Zhao J, et al. Metronomic
docetaxel chemotherapy inhibits angiogenesis
and tumor growth in a gastric cancer model.
Cancer Chemother Pharmacol 2011;68:879–87.
Kerbel RS, Kamen BA. The anti-angiogenic basis
of metronomic chemotherapy. Nat Rev Cancer
2004;4:423–36.
Bertolini F, Paul S, Mancuso P, et al. Maximum
tolerable dose and low-dose metronomic
chemotherapy have opposite effects on the
mobilization and viability of circulating
endothelial progenitor cells. Cancer Res 2003;63:
4342–6.
Rubie H, Coze C, Plantaz D, et al. Localised and
unresectable neuroblastoma in infants: excellent
outcome with low-dose primary chemotherapy.
Br J Cancer 2003;89:1605–9.
Buckstein R, Kerbel RS, Shaked Y, et al. HighDose celecoxib and metronomic "low-dose"
cyclophosphamide is an effective and safe
therapy in patients with relapsed and refractory
aggressive histology non-Hodgkin’s lymphoma.
Clin Cancer Res 2006;12:5190–8.
Young SD, Whissell M, Noble JC, et al. Phase II
clinical trial results involving treatment with
low-dose daily oral cyclophosphamide, weekly
vinblastine, and rofecoxib in patients with
advanced solid tumors. Clin Cancer Res 2006;12:
3092–8.
Colleoni M, Rocca A, Sandri MT, et al. Lowdose oral methotrexate and cyclophosphamide
in metastatic breast cancer: antitumor activity
and correlation with vascular endothelial growth
factor levels. Ann Oncol 2002;13:
73–80.
Zou W, Restifo NP. T(H)17 cells in tumour
immunity and immunotherapy. Nat Rev
Immunol 2005;10:248–56.
Ghiringhelli F, Menard C, Puig PE, et al.
Metronomic cyclophosphamide regimen
selectively depletes CD4þCD25þ regulatory T
cells and restores T and NK effector functions
in end stage cancer patients. Cancer Immunol
Immunother 2007;56:641–8.
Ghiringhelli F, Larmonier N, Schmitt E, et al.
CD4þCD25þ regulatory T cells suppress tumor
immunity but are sensitive to cyclophosphamide
which allows immunotherapy of established
tumors to be curative. Eur J Immunol 2004;34:
336–44.
Liu JY, Wu Y, Zhang XS, et al. Single
administration of low dose cyclophosphamide
augments the antitumor effect of dendritic cell
62.
63.
64.
65.
66.
67.
68.
69.
70.
71.
72.
73.
74.
75.
76.
vaccine. Cancer Immunol Immunother 2007;56:
1597–604.
Lutsiak ME, Semnani RT, De Pascalis R, et al.
Inhibition of CD4(þ)25þ T regulatory cell
function implicated in enhanced immune
response by low-dose cyclophosphamide. Blood
2005;105:2862–8.
Taieb J, Chaput N, Schartz N, et al.
Chemoimmunotherapy of tumors:
cyclophosphamide synergizes with exosome
based vaccines. J Immunol 2006;176:2722–9.
Tongu M, Harashima N, Yamada T, et al.
Immunogenic chemotherapy with
cyclophosphamide and doxorubicin against
established murine carcinoma. Cancer Immunol
Immunother 2010;59:769–77.
Veltman JD, Lambers ME, van Nimwegen M,
et al. Low-dose cyclophosphamide synergizes
with dendritic cell-based immunotherapy in
antitumor activity. J Biomed Biotechnol 2010;
2010:798467.
Radojcic V, Bezak KB, Skarica M, et al.
Cyclophosphamide resets dendritic cell
homeostasis and enhances antitumor immunity
through effects that extend beyond regulatory T
cell elimination. Cancer Immunol Immunother
2010;59:137–48.
Moschella F, Proietti E, Capone I, et al.
Combination strategies for enhancing the
efficacy of immunotherapy in cancer patients.
Ann N Y Acad Sci 2010;1194:169–78.
del Hoyo GM, Martin P, Vargas HH, et al.
Characterization of a common precursor
population for dendritic cells. Nature 2002;415:
1043–7.
Yoneyama H, Matsuno K, Zhang Y, et al.
Regulation by chemokines of circulating
dendritic cell precursors, and the formation of
portal tract-associated lymphoid tissue, in a
granulomatous liver disease. J Exp Med 2001;
193:35–49.
Kaneno R, Shurin GV, Tourkova IL, et al.
Chemomodulation of human dendritic cell
function by antineoplastic agents in low
noncytotoxic concentrations. J Transl Med 2009;
7:58.
Garcia AA, Hirte H, Fleming G, et al. Phase II
clinical trial of bevacizumab and low-dose
metronomic oral cyclophosphamide in recurrent
ovarian cancer: a trial of the California, Chicago,
and Princess Margaret Hospital phase II
consortia. J Clin Oncol 2008;26:76–82.
Malvicini M, Rizzo M, Alaniz L, et al. A novel
synergistic combination of cyclophosphamide
and gene transfer of interleukin-12 eradicates
colorectal carcinoma in mice. Clin Cancer Res
2009;15:7256–65.
Ercolini AM, Ladle BH, Manning EA, et al.
Recruitment of latent pools of high-avidity
CD8(þ) T cells to the antitumor immune
response. J Exp Med 2005;201:1591–602.
North RJ. Cyclophosphamide-facilitated adoptive
immunotherapy of an established tumor
depends on elimination of tumor-induced
suppressor T cells. J Exp Med 1982;
155:1063–74.
Demicheli R, Valagussa P, Bonadonna G. Does
surgery modify growth kinetics of breast cancer
micrometastases? Br J Cancer 2001;85:490–2.
Emmenegger U, Morton GC, Francia G, et al.
Low-dose metronomic daily cyclophosphamide
and weekly tirapazamine: a well-tolerated
combination regimen with enhanced efficacy
Mini Review
2477
Nars and Kaneno
2478
77.
78.
Mini Review
79.
80.
81.
82.
83.
84.
85.
that exploits tumor hypoxia. Cancer Res 2006;
66:1664–74.
Yu JL, Rak JW, Coomber BL, et al. Effect of p53
status on tumor response to
antiangiogenic therapy. Science 2002;295:
1526–8.
Ogawa Y, Ishikawa T, Chung SH, et al. Oral
UFT and cyclophosphamide combination
chemotherapy for metastatic breast cancer.
Anticancer Res 2003;23:3453–7.
Zhong H, Han B, Tourkova IL, et al. Low-dose
paclitaxel prior to intratumoral dendritic cell
vaccine modulates intratumoral cytokine
network and lung cancer growth. Clin Cancer
Res 2007;13:5455–62.
Ramakrishnan R, Assudani D, Nagaraj S, et al.
Chemotherapy enhances tumor cell
susceptibility to CTL-mediated killing during
cancer immunotherapy in mice. J Clin Invest
2010;120:1111–24.
He Q, Li J, Yin W, et al. Low-dose paclitaxel
enhances the anti-tumor efficacy of GM-CSF
surface-modified whole-tumor-cell vaccine in
mouse model of prostate cancer. Cancer
Immunol Immunother 2011;
60:715–30.
Jiang H, Tao W, Zhang M, et al. Low-dose
metronomic paclitaxel chemotherapy suppresses
breast tumors and metastases in mice. Cancer
Invest 2010;28:74–84.
Ko HJ, Kim YJ, Kim YS, et al. A combination of
chemoimmunotherapies can efficiently break
self-tolerance and induce antitumor immunity
in a tolerogenic murine tumor model. Cancer
Res 2007;67:7477–86.
Suzuki E, Kapoor V, Jassar AS, et al.
Gemcitabine selectively eliminates splenic Gr1þ/CD11bþ myeloid suppressor cells in tumorbearing animals and enhances antitumor
immune activity. Clin Cancer Res 2005;11:
6713–21.
Correale P, Cerretani D, Remondo C, et al. A
novel metronomic chemotherapy regimen of
weekly platinum and daily oral etoposide in
high-risk non-small cell lung cancer patients.
Oncol Rep 2006;16:133–40.
Immunomodulation by low dose chemotherapy
86.
87.
88.
89.
90.
91.
92.
93.
94.
95.
Correale P, Remondo C, Carbone SF, et al.
Dose/dense metronomic chemotherapy with
fractioned cisplatin and oral daily etoposide
enhances the anti-angiogenic effects of
bevacizumab and has strong antitumor activity
in advanced non-small-cell-lung cancer patients.
Cancer Biol Ther 2010;9:685–93.
Correale P, Botta C, Basile A, et al. Phase II trial
of bevacizumab and dose/dense chemotherapy
with cisplatin and metronomic daily oral
etoposide in advanced non-small-cell-lung
cancer patients. Cancer Biol Ther 2011;12:112–8.
Correale P, Botta C, Cusi MG, et al. Cetuximab
þ/ chemotherapy enhances dendritic cellmediated phagocytosis of colon cancer cells and
ignites a highly efficient colon cancer antigenspecific cytotoxic T-cell response in vitro.
Int J Cancer 2012;130:1577–89.
Lake RA, Robinson BW. Immunotherapy and
chemotherapy-a practical partnership. Nat Rev
Cancer 2005;5:397–405.
Shurin GV, Tourkova IL, Kaneno R, et al.
Chemotherapeutic agents in noncytotoxic
concentrations increase antigen presentation by
dendritic cells via an IL-12-dependent
mechanism. J Immunol 2009;183:137–44.
Curtsinger JM, Lins DC, Mescher MF. Signal 3
determines tolerance versus full activation of
naive CD8 T cells: dissociating proliferation and
development of effector function. J Exp Med
2003;197:1141–51.
Mescher MF, Curtsinger JM, Agarwal P, et al.
Signals required for programming effector and
memory development by CD8þ T cells.
Immunol Rev 2006;211:81–92.
Tong AW, Papayoti MH, Netto G, et al.
Growth-inhibitory effects of CD40 ligand
(CD154) and its endogenous expression in
human breast cancer. Clin Cancer Res 2001;7:
691–703.
Shurin MR, Yurkovetsky ZR, Tourkova IL, et al.
Inhibition of CD40 expression and CD40mediated dendritic cell function by tumorderived IL-10. Int J Cancer 2002;101:61–8.
Pinzon-Charry A, Schmidt CW, Lopez JA. The
key role of CD40 ligand in overcoming tumor-
96.
97.
98.
99.
100.
101.
102.
103.
104.
105.
induced dendritic cell dysfunction. Breast
Cancer Res 2006;8:402.
Pirtskhalaishvili G, Shurin GV, Esche C, et al.
Cytokine-mediated protection of human
dendritic cells from prostate cancer-induced
apoptosis is regulated by the Bcl-2 family of
proteins. Br J Cancer 2000;
83:506–13.
Esche C, Gambotto A, Satoh Y, et al. CD154
inhibits tumor-induced apoptosis in dendritic
cells and tumor growth. Eur J Immunol 1999;29:
2148–55.
Sevko A, Kremer V, Falk C, et al. Application of
paclitaxel in low non-cytotoxic doses supports
vaccination with melanoma antigens in normal
mice. J Immunotoxicol, 2012;9:275–81.
Michels T, Shurin GV, Naiditch H, et al.
Paclitaxel promotes differentiation of
myeloid-derived suppressor cells into dendritic
cells in vitro in a TLR4-independent
manner. J Immunotoxicol, 2012;9:
292–300.
Kaneno R, Shurin GV, Kaneno FM, et al.
Chemotherapeutic agents in low noncytotoxic
concentrations increase immunogenicity of
human colon cancer cells. Cell Oncol (Dordr)
2011;34:97–106.
Haynes NM, van der Most RG, Lake RA, et al.
Immunogenic anti-cancer chemotherapy as an
emerging concept. Curr Opin Immunol 2008;20:
545–57.
Tesniere A, Apetoh L, Ghiringhelli F, et al.
Immunogenic cancer cell death: a key-lock
paradigm. Curr Opin Immunol 2008;
20:504–11.
Obeid M, Tesniere A, Ghiringhelli F, et al.
Calreticulin exposure dictates the
immunogenicity of cancer cell death. Nat Med
2007;13:54–61.
Melief CJ. Cancer immunotherapy by dendritic
cells. Immunity 2008;29:372–83.
Tanaka H, Matsushima H, Nishibu A, et al.
Dual therapeutic efficacy of vinblastine as a
unique chemotherapeutic agent capable of
inducing dendritic cell maturation. Cancer Res
2009;69:6987–94.
C 2012 UICC
Int. J. Cancer: 132, 2471–2478 (2013) V