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NEOPLASIA
From the Genetics Institute, Wyeth-Ayerst Research,
Cambridge, MA.
Major mechanisms underlying poor immune responses to autologous
tumor-associated antigens are overwhelming tumor kinetics and
the absence of effective T-cell costimulation by antigen-presenting cells. To address these issues, leukemia and lymphoma mice were treated with the combination of chemotherapy and systemic
immunotherapy with recombinant soluble murine B7-immunoglobulin G
(IgG) molecules. In this report, 3 murine models were used, a
radiation-induced SJL acute myeloid leukemia, a transplantable
spontaneous SJL lymphoma, and the C57BL/6 EL-4 thymic lymphoma. Various
treatment modalities were evaluated: single treatments with
either B7-IgG or chemotherapy as well as combination therapies.
The results demonstrate the following: (1) in all tumor models, the
combination of chemotherapy and soluble B7-IgGs is more potent than
either therapy alone, leading to cure of tumor-bearing animals; (2) the
therapeutic responses are T-cell-dependent, because combined therapy
is not efficacious in severe combined immunodeficient mice; (3) the
rejection of tumor cells leads to the development of tumor-specific
immunity, because cured mice are immune to the rejected tumor but not
to a different syngeneic tumor; and (4) 51Cr release assays
show that rejection of tumor cells leads to the development of very
potent tumor-specific cytotoxic T-lymphocyte activity. On the basis of
these results, it is proposed that chemotherapy-mediated tumor
reduction, together with consequent augmented tumor-antigen presentation to activated T cells, are primary mechanisms leading to
curative responses. The safety profile of the B7-IgG fusion proteins
and their synergy with chemotherapy strongly suggest that the
combination regimen is a promising strategy in cancer treatment.
(Blood. 2001;97:2420-2426) Major recent discoveries that have drastically
modified the nature of T-cell-directed immunotherapy in cancer are the
cloning of several tumor-associated antigens (TAAs) that elicit
autologous cytotoxic T-cell responses1,2 and the discovery
of new molecules and pathways involved in T-cell activation and
costimulation.3,4 This knowledge has guided the design of
numerous preclinical and clinical studies that, in turn, have generated
remarkable insight into the mechanisms controlling host responses to
cancer cells. Thus, it is now accepted that few spontaneous tumors are
immunogenic, but most, if not all, are antigenic.5,6
However, what ultimately determines the outcome of an endogenous
antigen encounter is the context in which that particular antigen is
presented to T cells.7,8 In the absence of danger signals
that accompany tissue destruction and inflammation (typically observed
during viral infection), the outcome is immune
ignorance.9,10 At the level of antigen-presenting cell
(APC)-T-cell interactions, the local danger signals translate primarily into the up-regulation of T-cell costimulatory signals provided by APCs at the time of antigen presentation.
Costimulation is defined as a signal necessary for optimal T-cell
activation and survival delivered to T cells along with the T-cell
receptor (TCR) signal, provided by APCs, ie, activated B cells,
macrophages, and dendritic cells.11,12 Over the years, the
CD28/B7 T-cell costimulatory pathway has emerged as the key regulator
of T-cell responses. The signal involves the interaction of the T-cell
surface antigen CD28 with the members of the B7 family molecules B7.1
(CD80) and B7.2 (CD86) expressed on APCs.13,14 Following a
TCR-mediated signal, ligation of CD28 results in up-regulation of the
interleukin 2 (IL-2)-receptor, increased IL-2 messenger RNA
transcription, T-cell proliferation, cytokine secretion, and up-regulation of several T-cell-activation-related
molecules.15 In contrast to CD28-mediated costimulation,
the CD28 homologue cytotoxic T-lymphocye antigen-4 (CTLA-4, CD152)
delivers a down-regulatory signal.16,17 In this context,
CTLA-4-mediated immune regulation appears to be critical for the
maintenance of immunological homeostasis.18
The discovery of the B7-CD28/CTLA-4 pathway led to the speculation that
lack of T-cell costimulation could be an important mechanism conferring
low immunogenicity, even to tumors expressing major histocompatibility
complex (MHC) molecules, which are able to present tumor antigens to T
cells.19,20 The hypothesis that introduction of B7 genes
into tumors might result in effective tumor immunogenicity has been
demonstrated in several tumor models. The pioneering work of Chen et
al21 and Townsend and Allison22 showed that
genetic modification of melanoma cells to express B7.1 led to
reduced tumorigenicity of the modified cells and to the development of
tumor-specific immunity. Subsequent studies demonstrated the efficacy
of B7.1-based tumor vaccines in myeloid leukemia
models.23,24 A striking observation that emerged from these studies was the lack of efficacy against bulk disease; in this
context, B7.1-based tumor vaccines were efficacious only when given in
early stages of the disease. It has been proposed that this is simply
due to drastically increased replication rates of tumor cells that may
preclude the opportunity for developing an adequate immune
response.25,26
In an attempt to overcome the practical problems currently related to
genetic modification of patients' tumor cells, we have developed
soluble B7-immunoglobulin G (IgG) (B7.1-IgG and B7.2-IgG) fusion
proteins and have recently shown their efficacy as single therapy in
several murine solid-tumor models.27 In this report, we
tested the efficacy of soluble B7-IgG in murine systemic leukemia and
lymphoma models. Our results show that in poorly immunogenic and
aggressive systemic tumor models, single therapy with either B7-IgG
fusion proteins or cytoreductive chemotherapy has variable efficacy,
but neither therapy alone is able to cure tumor-bearing animals.
However, when both systemic immunotherapy and chemotherapy are
combined, leukemia and lymphoma tumors are rejected and treated mice
are cured.
Mice
Tumor models
Murine recombinant B7-IgG fusion proteins The development and purification of soluble B7.1- and B7.2-IgG fusion proteins have been previously described.27 Briefly, complementary DNA encoding the signal and the extracellular domains of murine B7.1 or B7.2 were joined to the genomic DNA encoding the hinge-CH2-CH3 domains derived from a murine IgG2a antibody. B7-IgG molecules are dimeric and can bind to CD28 and/or CTLA-4 on T cells and to Fc receptors (FcRs) on macrophages and APCs. In some experiments, either mutated B7.2-IgG fusion protein (the IgG2a region is mutated to ablate binding to Fc receptor I [FcRI]and complement C1b) or
control isotype murine IgG2a was used.
Chemotherapy protocols Leukemia protocol was as follows: cytarabine (Cytosar-U, Pharmacia & Upjohn, Kalamazoo, MI), 2 consecutive intraperitoneal (IP) injections of 200 mg/kg, 6 hours apart, on days 7, 14, and 21; doxorubicin hydrochloride (Adriamycin RDF, Pharmacia & Upjohn), 1 IP injection of 6 mg/kg on days 7 and 14. Lymphoma protocol was as follows: cyclophosphamide monohydrate (Sigma, St Louis, MO), 1 IP injection of 100 mg/kg on days 7 and 21; doxorubicin hydrochloride, 1 IP injection of 6 mg/kg on days 7 and 21. The choice of the compounds was based on their use in clinical leukemia and lymphoma chemotherapy protocols.Combination therapy Mice were injected with live tumor cells on day 0 and subsequently treated with chemotherapy and one injection of 100 µg B7.2-IgG (or 50 µg of each B7.2- and B7.1-IgG) on days 7, 14, 21, and 28. B7-IgG was diluted in 200 µL of sterile saline and administered by subcutaneous injections at the described doses and schedules. Tumor size in EL-4 and survival of mice in AML and Ly were monitored twice weekly. Except for survival experiments, mice were killed when tumors reached a size of approximately 400 to 600 mm2.Proliferation assays Spleens were harvested from nontreated leukemic (AML) mice and from mice that had received 1 (day 7) or 2 (days 7 and 14) courses of chemotherapy (AML plus chemotherapy). Single-cell suspensions were prepared as previously described. Cells were cultured at 2 × 105 cells per well in flat-bottomed 96-well plates coated with a suboptimal dose (100 ng/mL) of anti-CD3 monoclonal antibody (mAb) 145-2C11(Pharmingen, San Diego, CA) and increasing amounts of plate-bound B7.2-IgG. Response to costimulation with anti-CD3 plus anti-CD28 (1 µg/mL) was used as positive control. Proliferation of responder cells was measured after 72 hours by the incorporation of 3H thymidine (1 µCi per well) for the last 18 hours of incubation. In tumor cell proliferation assays, 106 tumor cells per milliliter (2 × 105 cells per well in U-bottomed 96-well plates) were cultured with 10 µg/mL of soluble B7.1- or B7.2-IgG or control IgG, and their proliferation was measured 72 hours after culture initiation.Murine IL-2 enzyme-linked immunosorbent assay Levels of murine IL-2 in culture supernatants were determined by a sandwich enzyme-linked immunosorbent assay that used specific antimurine mAbs for capture and detection (Endogen, Woburn, MA). The sensitivity of the assay is 5 pg/mL.Immunostaining and flow cytometry analysis Splenocytes were isolated from AML and AML-plus-chemotherapy mice and were stained as previously described.24 The following mAbs (Pharmingen) were used for flow cytometry studies: Gr-1+ (RB6-8C5), CD3e (145-2C11), CD4 (L3T4), CD8a (53-67).51Cr release cytotoxic T-lymphocyte assays Spleens were collected from mice 11 weeks after EL-4 tumor inoculation/rejection, and single-cell suspensions were prepared. Splenocytes (5 × 106) were cocultured with irradiated (7335 cGy) EL-4 cells (1 × 105) in 2 mL complete RPMI per well of a 24-well tissue-culture plate (Costar, Cambridge, MA). At 6 days later, splenocytes were harvested and used as effector cells in cytotoxic T-lymphocyte (CTL) assays. EL-4 cells or control AML cells (2 × 106) were labeled with 200 µCi 51Cr (New England Nuclear, Boston, MA) for 90 minutes, washed twice, and used as targets (5000 per well) in the CTL assays. The standard 4-hour CTL assays were set up with various effector-to-target (E/T) ratios as previously described.Statistical analysis Most individual experiments consisted of 10 mice per treatment group. The data analyzed represent the results of 2 or 3 individual experiments. The statistical survival analysis was performed by means of the standard Mantel-Cox log-rank test. Cytokine values and proliferation results are the mean ± SD. The statistical significance between various groups was analyzed by Student t test.
Effects of systemic chemotherapy In an attempt to mimic clinical situations as closely as possible, we used chemotherapy regimens consisting of drugs that are major components of leukemia and lymphoma clinical protocols. Because most cytoreductive drugs target dividing cells, it is conceivable that any proliferating cells of the immune system would also be affected. Therefore, we first determined what the in vivo effects of chemotherapy were on both tumor cells and T cells. We treated leukemic mice on days 7 and 14 with Ara-C plus doxorubicin (AML plus chemotherapy). On day 14 or 21 (1 week after the first or second dose of chemotherapy), AML-plus-chemotherapy mice and control AML mice were killed, and their splenocytes were used for in vitro studies. Immunostaining and flow cytometric analysis on day 14 showed that after one course of chemotherapy, AML-plus-chemotherapy spleen had decreased numbers of Gr-1+ cells (marker for the leukemic cells)24 and increased numbers of T cells as compared with AML spleen (data not shown). The effect of chemotherapy was more prominent in day-21 flow cytometric analysis. As shown in Figure 1A, there was a dramatic decrease in the percentage of Gr-1+ cells in AML-plus-chemotherapy spleen as compared with AML (28% vs 87.2%), whereas the percentage of CD3, CD4, and CD8 T cells was significantly increased in AML-plus-chemotherapy spleen (29.1% vs 10.3%, 16.3% vs 2.4%, and 7.2% vs 0.94%, respectively).
We next determined the proliferative and cytokine response of AML-plus-chemotherapy splenocytes to in vitro B7.2-IgG costimulation. On day 14, no differences were observed between AML-plus-chemotherapy and AML splenocytes (Figure 1B), and both appeared to proliferate more vigorously with B7-IgG costimulation than with costimulation by anti-CD28 mAb. Furthermore, as shown in Figure 1C, IL-2 secretion in response to B7-IgG costimulation was comparable to stimulation through CD28. (The IL-2 levels in response to 5 µg/mL AML-plus-chemotherapy culture presented in Figure 1C were not observed in any of our other, similar in vitro costimulation experiments. These data are included because they are part of the series of other parameter measurements in the same experimental system). Overall, no major differences were observed between AML-plus-chemotherapy and AML splenocytes, indicating that this chemotherapy regimen did not cause detectable immunosuppression of the leukemic mice with regard to thymidine uptake and cytokine secretion. The day-21 proliferation and IL-2 level assays were performed only on AML-plus-chemotherapy splenocytes (as shown in Figure 1A, day-21 AML spleens were heavily infiltrated with Gr-1+ cells), and the results were comparable to those from day-14 assays (data not shown). Collectively, these results demonstrate that treatment of murine AML with doxorubicin and Ara-C significantly reduces the leukemic burden, without having a detrimental effect on in vitro T-cell proliferation and IL-2 secretion in response to B7.2-IgG costimulation. In vitro effects of B7.2-IgG on tumor cells Soluble B7.2-IgG fusion protein can potentially bind on CD28/CTLA-4 and FcRs on cells. Because EL-4 cells express CD28 and all 3 tumor cell types express FcRs, we investigated what the in
vitro direct effects of soluble B7.2-IgG were on tumor cells. Tumor
cells were cultured as described in "Materials and methods," and
their immunophenotype (fluorescence-activated cell sorting analysis),
viability (trypan-blue exclusion), and proliferative response
(3H thymidine uptake) were examined. In all types of
experiments, no significant differences were observed among the media,
IgG, or B7-IgG cultures (data not shown), indicating that there is no
direct effect of soluble B7.1- or B7.2-IgG on tumor cells with regard
to their immunophenotype, viability, and proliferative profile.
Combination therapy leads to cure of leukemia and lymphoma mice Single and combined treatment modalities (with B7.2-IgG, B7.1/B7.2-IgG, mutated form of B7.2-IgG, or control IgG) were evaluated in each tumor model, and no statistically significant differences were observed with the use of different sets of circumstances. Each experiment was repeated 2 to 3 times with similar results, and data from one representative experiment per model will be presented in this report. In the experiment shown in Figure 2A, single B7.2-IgG or control IgG treatment in the AML model had no effect. In most experiments, AML mice treated with single chemotherapy had 1 to 2 weeks' prolonged survival, as compared with untreated animals (P < .05), but eventually all mice developed lethal leukemia. However, when both treatments were combined, 50% of the mice were cured (Figure 2A).
In the EL-4 model, single therapy with either B7.2-IgG or chemotherapy
alone had minimal or no effect. Treatment with B7.2-IgG or the
mutated form of B7.2-IgG (which does not bind high-affinity Fc In the SJL Ly model, single therapy with fusion proteins showed no therapeutic effect (Figure 2C). Chemotherapy alone significantly prolonged the survival of treated mice (P < .01), but no cures were observed. In the same experiment, B7.2-IgG plus chemotherapy led to 90% cure, and the combination of B7-IgGs plus chemotherapy to 70% cure. Collectively, these results demonstrate that, in all models tested in this study, the combination of B7-IgG fusion proteins with chemotherapy leads to cure of leukemia- and lymphoma-bearing mice. Therapeutic responses to combination therapy are T-cell dependent It has been previously shown in tumor models that B7-IgG-mediated curative responses are CD8+ T-cell-dependent.27 To examine the role of T cells in the therapeutic responses to combination therapy, we evaluated single therapies (B7-IgG or chemotherapy) and combination therapy in T- and B-cell-deficient (SCID) mice. C57BL/6 SCID mice bearing EL-4 tumors were treated with B7.2-IgG, chemotherapy alone, or the combination of both. The tumors grew more rapidly in SCID mice than in normal C57BL/6 mice, and by day 28 after tumor inoculation, all mice had succumbed to lethal tumors. As shown in Figure 3, combination therapy had no therapeutic effect in SCID mice, clearly indicating the indispensable role of T cells for the efficacy of the combination regimen. Combination experiments in C57BL/6 SCID mice were also pursued in another systemic leukemia model (C1498 myeloid leukemia model), and similar results were observed (K.D.-J., unpublished data, March 2000).
Combination therapy leads to long-lasting tumor-specific immunity One of the main goals of combining cytoreductive chemotherapy with immunotherapy is to activate the adaptive immune system and thus trigger the development of effector and memory cytotoxic T cells. Whereas effector T cells in this setting may have a beneficial role in eliminating residual tumor cells that have escaped chemotherapy, long-term disease-free survival can be achieved only when potential tumor relapses are under the critical control of memory cytotoxic T cells. Therefore, we sought to determine if combination therapy was able to support the development of antitumor memory T cells. AML mice that had been cured with combination therapy were challenged 4 months later with live AML cells. As shown in Figure 4A, all mice were immune to challenge and rejected the leukemic cells. At 2 months after the AML challenge and rejection, the same mice were inoculated with syngeneic lymphoma (Ly) cells. As shown in Figure 4B, the challenged mice developed lethal lymphoma at the same time as naive mice that had been inoculated with Ly cells. These findings demonstrate that the mechanisms mediating curative responses in combination therapy also lead to the development of long-lived, tumor-specific memory cells.
Combination therapy leads to long-lasting tumor-specific CTL activity To further characterize the antitumor memory responses of cured mice, we performed in vitro CTL assays. Spleens were harvested from mice that had been cured of EL-4 lymphoma (with B7.2-IgG plus chemotherapy or the combination of B7-IgG plus chemotherapy), and splenocytes were assayed for in vitro CTL activity. As shown in Figure 5, both groups of cured mice generated very potent cytolytic responses upon stimulation with EL-4 cells. The response was EL-4-specific (H-2b) because the same cells did not lyse alloantigen-presenting AML (H-2s) cells.
In this report, we demonstrate that the addition of soluble B7.2-IgG to conventional leukemia and lymphoma chemotherapy regimens has remarkable synergistic effects in murine leukemia and lymphoma models, leading to curative T-cell-dependent antitumor responses. The establishment of long-lived tumor-specific memory T cells was confirmed with in vivo challenge experiments and in vitro CTL assays. Various mechanisms may contribute to the synergy observed in combination therapy, as opposed to the limited efficacy of single therapies in our studies. Chemotherapy alone, as shown in the AML model, significantly reduces the leukemic burden, but cannot eradicate minimal residual disease. Apparently, this is the reason chemotherapy-treated animals finally succumb to lethal leukemia, a situation resembling the relatively short duration of remission observed in AML patients with partial remission to induction chemotherapy.28,29 In addition to tumor-burden reduction, cytoreductive chemotherapy generates a plethora of TAAs, which, as is the case with all non-self-antigens, are eventually expressed in the context of MHC molecules on APCs and can potentially initiate antigen-specific T-cell activation. However, chemotherapy overall fails to generate clinically overt antitumor memory responses, primarily for 2 reasons: first, it temporarily reduces the T-cell pool by targeting the relatively low numbers of proliferating T cells, and secondly, the sudden availability of abundant tumor antigens is probably not accompanied by signals required for the maturation process of immature dendritic cells (DCs), the most potent APCs and initiators of immunity.30,31 In the presence of a maturation signal, DCs express higher levels of costimulatory and MHC molecules and can then activate resting T cells.32 In our studies, neither tumor reduction and increased TAA presentation (chemotherapy) nor T-cell activation (B7-IgG) alone can cure leukemia and lymphoma mice. The efficacy of the combination regimen suggests that B7-IgG soluble molecules are probably strengthening APC-T-cell interactions during antigen presentation, which in combination with chemotherapy-mediated tumor reduction can lead to curative immune responses. How does soluble B7-IgG fusion protein work? Because of the
nature of the fusion molecule, there is little doubt that in vivo it
will bind to both CD28/CTLA-4- and Fc The indispensable role of T cells in combination therapy was confirmed in studies with SCID mice, in which all experimental groups, with or without treatment, rapidly developed lethal tumors. The relatively complex nature of combination therapy has made us hesitant to use additional in vivo compounds for CD4+ and CD8+ T-cell depletion and determination of each subset's role in combination therapy. We plan, however, on specifically addressing the role of CD4+ and CD8+ cells in future work by (1) in vivo selective depletion of CD4/CD8 cells at the time of rechallenge, (2) adaptive transfer experiments, and (3) T-cell depletion of long-term survivors and determination of whether there is tumor regrowth at that time. In addition to CD28/CTLA-4, B7.2-IgG also binds in vitro (and
potentially in vivo) to Fc A critical issue in the combination regimen is the appropriate timing for the immunoadjuvant. It is well known that chemotherapy induces peripheral leukopenia whose severity and duration depend on the type and dose of chemotherapeutics.40 Whereas high-dose, myeloablative regimens are accompanied by prolonged peripheral leukopenia, conventional chemotherapy is related primarily to neutropenia and, to a lesser extent, to lymphopenia. Several studies in humans have shown that among lymphocyte subsets, B cells are the cells most affected by chemotherapy, followed by CD4+ T cells, with CD8+ T cells remaining relatively well preserved.41-44 Interestingly, the remaining clonogenic T lymphocytes derived from acute leukemia patients with therapy-induced leukopenia have shown a broad cytokine response to in vitro activation.45 We have previously shown in the SJL AML model that absolute peripheral lymphocyte numbers return to normal within a week after chemotherapy with cytarabine and doxorubicin.26 On the basis of these findings, we reasoned that the interval between injections of B7-IgG should be 1 week or longer. Because 2 of the B7-IgG injections in the lymphoma models and 3 in the AML model were given on the same day with chemotherapy, it is conceivable that a percentage of T cells responding to B7/CD28 interactions by proliferation would be affected by chemotherapy. However, the favorable clinical outcome of the combination regimen in our studies suggests that a significant number of B7-IgG-activated T cells can nevertheless resist chemotherapy and eventually become long-lived antitumor memory cells. A major potential advantage for the use of B7.2-IgG as an
adjuvant to chemotherapy is its expected high safety profile. The protein has not shown any in vivo toxicity, even when injections of 500 µg were given to mice (unpublished results, May 1999). Studies in the MethA tumor model (which has reproducibly shown therapeutic responses to single B7.2-IgG treatment) have shown that the
efficacy of the fusion protein is not ablated in IFN- The combination of immunotherapy with chemotherapy is an emerging form of cancer treatment. With the addition of an immune-boosting agent that, in principle, forces "provoked" immunity, conventional cancer therapy could conceivably be made more effective without increasing its toxicity.46 This may be manifested in greater durability of response rather than absolute clinical response rate. It is anticipated that individual immunomodulatory compounds will not synergize with all cytotoxic drugs, owing to differential immunosuppressive effects, and at present it is a great challenge to identify successful combinations in preclinical tumor models. The chemotherapy regimens used in this report apply beyond leukemia and lymphoma, since the combination of anthracyclines and cyclophosphamide is broadly used in the clinic and is one of the most widely used regimens in breast cancer treatment.47 The safety profile of combining B7.2-IgG with chemotherapy in preclinical tumor models, together with the potent therapeutic effect, has directed our efforts toward the development of strategies for clinical evaluation.
We thank Lori Block and Terri Haire for technical help and Drs Stan Wolf and Frank Borriello for critical review of the manuscript.
Submitted July 13, 2000; accepted December 8, 2000.
All authors were employed by Genetics Institute at the time that this study was conducted.
The publication costs of this article were defrayed in part by page charge payment. Therefore, and solely to indicate this fact, this article is hereby marked "advertisement" in accordance with 18 U.S.C. section 1734.
Reprints: K. Dunussi-Joannopoulos, Genetics Institute, 1 Burtt Rd, Andover, MA 01810.
1. Boon T, Cerottini JC, Van den Eynde B, van der Bruggen P, Van Pel A. Tumor-antigens recognized by T lymphocytes. Annu Rev Immunol. 1994;12:337-365[CrossRef][Medline] [Order article via Infotrieve]. 2. Robbins PF, Kawakami Y. Human tumor antigens recognized by T cells. Curr Opin Immunol. 1996;5:628-636. 3. June CH, Ledbetter JA, Linsley PS, Thompson CB. Role of the CD28 receptor in T-cell activation. Immunol Today. 1990;11:211-216[CrossRef][Medline] [Order article via Infotrieve]. 4. Linsley PS, Ledbetter JA. The role of the CD28 receptor during T cell responses to antigen. Annu Rev Immunol. 1993;11:191-212[Medline] [Order article via Infotrieve] 5. Henderson RA, Finn OJ. Human tumor antigens are ready to fly. Adv Immunol. 1996;62:217-256[Medline] [Order article via Infotrieve]. 6. Sogn JA. Tumor immunology: the glass is half full. Immunity. 1998;9:757-763[CrossRef][Medline] [Order article via Infotrieve]. 7. Pardoll DM. Cancer vaccines. Nat Med. 1998;4:525-531[CrossRef][Medline] [Order article via Infotrieve]. 8. Marincola FM, Jaffee EM, Hicklin DJ, Ferrone S. Escape of human solid tumors from T-cell recognition: molecular mechanisms and functional significance. Adv Immunol. 2000;74:181-273[Medline] [Order article via Infotrieve]. 9. Matzinger P. Tolerance, danger, and the extended family. Annu Rev Immunol. 1994;12:991-1045[Medline] [Order article via Infotrieve].
10.
Fenton RG, Longo DL.
Danger versus tolerance: paradigms for future studies of tumor-specific cytotoxic T lymphocytes.
J Natl Cancer Inst.
1997;89:272-275 11. Mueller DL, Jenkins MK, Schwarz RH. Clonal expansion versus functional clonal inactivation: a costimulatory signalling pathway determines the outcome of T cell antigen receptor occupancy. Annu Rev Immunol. 1989;7:445-480[Medline] [Order article via Infotrieve]. 12. Schwartz RH. Acquisition of immunologic self-tolerance. Cell. 1989;57:1073-1081[CrossRef][Medline] [Order article via Infotrieve]. 13. June CH, Bluestone JA, Nadler LM, Thompson CB. The B7 and CD28 receptor families. Immunol Today. 1994;15:321-331[CrossRef][Medline] [Order article via Infotrieve]. 14. Lenschow DJ, Walunas TL, Bluestone JA. CD28/B7 system of T cell costimulation. Annu Rev Immunol. 1996;14:233-258[CrossRef][Medline] [Order article via Infotrieve]. 15. Boussiotis VA, Freeman GJ, Gribben JG, Nadler LM. The role of B7-1/B7-2:CD28/CTLA-4 pathways in the prevention of anergy, induction of productive immunity and down-regulation of the immune response. Immunol Rev. 1996;153:5-26[CrossRef][Medline] [Order article via Infotrieve]. 16. Allison JP. CD28-B7 interactions in T-cell activation. Curr Opin Immunol. 1994;6:414-419[CrossRef][Medline] [Order article via Infotrieve]. 17. Chambers CA, Krummel MF, Boitel B, et al. The role of CTLA-4 in the regulation and initiation of T-cell responses. Immunol Rev. 1996;153:27-46[CrossRef][Medline] [Order article via Infotrieve]
18.
Waterhouse P, Penninger JM, Timms E, et al.
Lymphoproliferative disorders with early lethality in mice deficient in CTLA-4.
Science.
1995;270:985-988 19. Chen L, Linsley PS, Hellström KE. Costimulation of T cells for tumor immunity. Immunol Today. 1993;14:483-486[CrossRef][Medline] [Order article via Infotrieve]. 20. Allison JP, Hurwitz AA, Leach DR. Manipulation of costimulatory signals to enhance antitumor T-cell responses. Curr Opin Immunol. 1995;7:682-686[CrossRef][Medline] [Order article via Infotrieve]. 21. Chen L, Ashe S, Brady WA, et al. Costimulation of antitumor immunity by the B7 counterreceptor for the T lymphocyte molecules CD28 and CTLA-4. Cell. 1992;71:1093-1102[CrossRef][Medline] [Order article via Infotrieve].
22.
Townsend SE, Allison JP.
Tumor rejection after direct costimualtion of CD8+ T cells by B7-transfected melanoma cells.
Science.
1993;259:368-370
23.
Matulonis UA, Dosiou C, Lamont C, et al.
Role of B7-1 in mediating an immune response to myeloid leukemia cells.
Blood.
1995;85:2507-2515
24.
Dunussi-Joannopoulos K, Weinstein HJ, Nickerson PW, et al.
Irradiated B7-1 transduced primary acute myelogenous leukemia (AML) cells can be used as therapeutic vaccines in murine AML.
Blood.
1996;87:2938-2946 25. Dranoff G, Mulligan RC. Gene transfer as cancer therapy. Adv Immunol. 1995;58:417-454[Medline] [Order article via Infotrieve].
26.
Dunussi-Joannopoulos K, Krenger W, Weinstein HJ, Ferrara JLM, Croop JM.
CD8+ T-cells activated during the course of murine acute myelogenous leukemia elicit therapeutic responses to late B7 vaccines after cytoreductive treatment.
Blood.
1997;89:2915-2924
27.
Sturmhoefel K, Lee K, Gray GS, et al.
Potent activity of soluble B7-IgG fusion proteins in therapy of established tumors and as vaccine adjuvant.
Cancer Res.
1999;59:4964-4972 28. Burnett AK, Eden OB. The treatment of acute leukaemia. Lancet. 1997;349:270-275[CrossRef][Medline] [Order article via Infotrieve].
29.
Estey EH, Shen Y, Thall PF.
Effect of time to complete remission on subsequent survival and disease-free survival time in AML, RAEB-t, and RAEB.
Blood.
2000;95:72-77 30. Gallucci S, Lolkema M, Matzinger P. Natural adjuvants: endogenous activators of dendritic cells. Nat Med. 1999;5:1249-1255[CrossRef][Medline] [Order article via Infotrieve].
31.
Sauter B, Albert ML, Francisco L, Larsson M, Somersan S, Bhardwaj N.
Consequences of cell death: exposure to necrotic tumor cells, but not primary tissue cells or apoptotic cells, induces the maturation of immunostimulatory dendritic cells [comment appears in J Exp Med. 2000;191:411-416].
J Exp Med.
2000;191:423-434 32. Banchereau J, Steinman RM. Dendritic cells and the control of immunity. Nature. 1998;392:245-252[CrossRef][Medline] [Order article via Infotrieve].
33.
Runyon K, Lee K, Leonard JP, Collins M, Dunussi-Joannopoulos K.
Fc 34. Freeman GJ, Lombard DB, Gimmi CD, et al. CTLA-4 and CD28 mRNA are coexpressed in most T cells after activation: expression of CTLA-4 and CD28 mRNA does not correlate with the pattern of lymphokine production. J Immunol. 1992;149:3795-3801[Abstract]. 35. Lindsten T, Lee KP, Harris ES, et al. Characterization of CTLA-4 structure and expression on human T cells. J Immunol. 1993;151:3489-3499[Abstract]. 36. Amigorena S, Salamero J, Davoust J, Fridman WH, Bonnerot C. Tyrosine-containing motif that transduces cell activation signals also determines internalization and antigen presentation via type III receptors for IgG. Nature. 1992;358:337-341[CrossRef][Medline] [Order article via Infotrieve]. 37. Bonnerot C, Amigorena S, Choquet D, Pavlovich R, Choukroun V, Fridman WH. Role of associated gamma-chain in tyrosine kinase activation via murine Fc gamma RIII. EMBO J. 1992;11:2747-2757[Medline] [Order article via Infotrieve].
38.
Amigorena S, Lankar D, Briken V, Gapin L, Viguier M, Bonnerot C.
Type II and III receptors for immunoglobulin G (IgG) control the presentation of different T cell epitopes from single IgG-complexed antigens.
J Exp Med.
1998;187:505-515 39. Gessner JE, Heiken H, Tamm A, Schmidt RE. The IgG Fc receptor family. Ann Hematol. 1998;76:231-248[CrossRef][Medline] [Order article via Infotrieve]. 40. Johnston EM, Crawford J. Hematopoietic growth factors in the reduction of chemotherapeutic toxicity. Semin Oncol. 1998;25:552-561[Medline] [Order article via Infotrieve]. 41. Petrini B, Wasserman J, Blomgren H, Baral E, Strender LE, Wallgren A. Blood lymphocyte subpopulations in breast cancer patients following post-operative adjuvant chemotherapy or radiotherapy. Clin Exp Immunol. 1979;38:361-365[Medline] [Order article via Infotrieve]. 42. Sewell HF, Halbert CF, Robins RA, Galvin A, Chan S, Blamey RW. Chemotherapy-induced differential changes in lymphocyte subsets and natural-killer-cell function in patients with advanced breast cancer. Int J Cancer. 1993;55:735-738[Medline] [Order article via Infotrieve].
43.
Mackall CL, Fleisher TA, Brown MR, et al.
Lymphocyte depletion during treatment with intensive chemotherapy for cancer.
Blood.
1994;84:2221-2228 44. Mackall CL. T-cell immunodeficiency following cytotoxic antineoplastic therapy: a review. Stem Cells. 2000;18:10-18[CrossRef][Medline] [Order article via Infotrieve]. 45. Bruserud O. Cellular immune responses in acute leukaemia patients with severe chemotherapy-induced leucopenia; characterization of the cytokine repertoire of clonogenic T cells. Cancer Immunol Immunother. 1998;46:221-228[CrossRef][Medline] [Order article via Infotrieve]. 46. Windbichler GH, Hausmaninger H, Stummvoll W, et al. Interferon-gamma in the first-line therapy of ovarian cancer: a randomized phase III trial. Br J Cancer. 2000;82:1138-1144[CrossRef][Medline] [Order article via Infotrieve]. 47. Dees EC, Kennedy MJ. Recent advances in systemic therapy for breast cancer. Curr Opin Oncol. 1998;10:517-522[Medline] [Order article via Infotrieve].
© 2001 by The American Society of Hematology.
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  N. Di Gaetano, E. Cittera, R. Nota, A. Vecchi, V. Grieco, E. Scanziani, M. Botto, M. Introna, and J. Golay Complement Activation Determines the Therapeutic Activity of Rituximab In Vivo J. Immunol., August 1, 2003; 171(3): 1581 - 1587. [Abstract] [Full Text] [PDF]
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 X. Huang, M. K. Wong, H. Yi, S. Watkins, A. D. Laird, S. F. Wolf, and E. Gorelik Combined Therapy of Local and Metastatic 4T1 Breast Tumor in Mice Using SU6668, an Inhibitor of Angiogenic Receptor Tyrosine Kinases, and the Immunostimulator B7.2-IgG Fusion Protein Cancer Res., October 15, 2002; 62(20): 5727 - 5735. [Abstract] [Full Text] [PDF]
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Top
Abstract
Introduction
)
received no treatment; the second group (AML plus chemotherapy, 
)
was treated with chemotherapy (chemo) as described in "Materials and
methods." (A) Flow cytometric analysis of AML and
AML-plus-chemotherapy splenocytes on day 21 (1 week after the second
dose of chemotherapy). In all panels, solid histograms represent the
expression of the indicated markers by splenocytes. (B) AML and
AML-plus-chemotherapy splenocytes were cultured at 2 ×105
cells per well in U-bottomed 96-well plates coated with suboptimal dose
(100 ng/mL) of anti-CD3 mAb 145-2C11 and the indicated amounts of
plate-bound B7.2-IgG. Response to costimulation with anti-CD3 plus
anti-CD28 (1 µg/mL) was used as positive control. Proliferation of
responder cells was measured after 72 hours by the incorporation of
3H thymidine (1 µCi per well) for the last 18 hours of
incubation. Results are representative of 2 independent experiments and
are shown as the mean ± SD of 8 cultures. (C) AML and
AML-plus-chemotherapy splenocytes were cultured as described in panel
B. IL-2 concentrations were determined in 24-hour tissue-culture
supernatants. Results are representative of 2 independent experiments
and are shown as the mean ± SD of 8 cultures.
) that had been cured with combination therapy
were challenged 4 months later with live 105 AML cells.
Naive SJL mice (