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The Journal of Immunology, 1999, 162: 4817-4823.
Copyright © 1999 by The American Association of Immunologists

Host B7-1 and B7-2 Costimulatory Molecules Contribute to the Eradication of B7-1-Transfected P815 Tumor Cells Via a CD8+ T Cell-Dependent Mechanism1

Ross N. La Motte2,*, Arlene H. Sharpe{dagger}, Jeffrey A. Bluestone3,{ddagger} and Margalit B. Mokyr3,4,*

* Department of Biochemistry and Molecular Biology, University of Illinois, Chicago, IL 60612; {dagger} Department of Pathology, Brigham and Women’s Hospital and Harvard Medical School, Boston, MA 02115; and {ddagger} Committee on Immunology, Department of Pathology, Ben May Institute for Cancer Research, University of Chicago, Chicago, IL 60637


   Abstract
Top
 Abstract
Introduction
 Materials and Methods
 Results
 Discussion
 References
 
B7-1 (CD80)-transfected P815 tumor cells were previously shown to elicit tumor-eradicating immunity that leads to the regression of B7-1+ P815 tumors after transient growth in normal syngeneic (DBA/2) mice. Here, we show that not only the B7-1 molecule but also the B7-2 (CD86) molecule contributed to the eradication of B7-1+ P815 tumors. The B7-1 molecule that contributed to the eradication of B7-1+ P815 tumors was expressed not only on the tumor cells but also on host APCs, including MAC-1+ cells. The B7-2 molecule that contributed to the eradication of B7-1+ P815 tumors was expressed only on host APCs, such as B220+ cells, and not on the tumor cells. In spite of the fact that B7-expressing host APCs contributed to the eradication of B7-1+ P815 tumors, only CD8+ T cells without help from CD4+ T cells were important for tumor eradication. Taken together, these findings indicate that in addition to the ability of B7-1-transfected tumor cells to stimulate CD8+ T cell-mediated tumor-eradicating immunity directly, such tumor cells can also stimulate CD8+ T cell-mediated tumor-eradicating immunity indirectly as a result of cross-priming through B7-expressing host APCs.


   Introduction
Top
 Abstract
 Introduction
Materials and Methods
 Results
 Discussion
 References
 
Tcell activation requires Ag-specific signals delivered through interaction between the TCR and the MHC/peptide complex, as well as Ag-independent costimulatory signals. In the case of CD4+ T cells, the costimulatory signals are delivered through interaction between CD28 expressed on the T cells and members of the B7 family expressed on APCs (1, 2, 3, 4). Activation of naive CD8+ T cells requires more costimulatory activity than activation of naive CD4+ T cells, and, in the presence of relatively low levels of B7 expression, CD8+ T cell activation is largely dependent on help from activated CD4+ T cells in the form of cytokines, such as IL-2 (5, 6). However, when relatively high levels of B7 are expressed on the APCs, CD8+ T cells can be activated directly and freed from the requirement for exogenous help by CD28-mediated autocrine IL-2 production (4, 6, 7).

Costimulation of T cells through CD28/B7 interaction was also shown to play an important role in eliciting tumor-eradicating immunity, and inadequate costimulation has been suggested to contribute to tumor progression in immunocompetent hosts (8, 9). Therefore, in an attempt to elicit a more powerful antitumor immunity, several groups of investigators have introduced the B7-1 and/or the B7-2 gene into tumor cells so as to turn them into efficient APCs (4, 10, 11). This approach was proven to be successful with a variety of tumor types and led to the rejection of the B7+ tumor cells by syngeneic normal mice with the acquisition of resistance to a subsequent challenge with unmodified parental tumor cells (9, 10, 11, 12, 13, 14, 15). The level of B7 expressed on the transfected tumor cells was quite high (11, 15, 16), and, in most cases, the tumor-eradicating immunity elicited by B7-transfected, MHC class I+ and class II- tumor cells was dependent on CD8+ T cells without help from CD4+ cells (10, 14, 16, 17, 18). These observations led to the conclusion that the B7-transfected tumor cells stimulate CD8+ T cells directly (4, 9).

The results presented in the current article confirm the importance of CD8+ T cells without help from CD4+ T cells for the eradication of B7-1-transfected, MHC class I+ and class II- tumor cells, with the P815 tumor model. In addition, our results extend these observations by illustrating that the CD8+ T cell-dependent tumor-eradicating immunity is generated not only as a result of directly activating the CD8+ T cells by stimulation with the B7-1+ P815 tumor cells, but also as a result of cross-priming via B7-1- and/or B7-2-expressing host APCs.


   Materials and Methods
Top
 Abstract
 Introduction
 Materials and Methods
Results
 Discussion
 References
 
Mice

Our studies employed primarily 7- to 12-wk-old female DBA/2 mice (H-2d) that were obtained from Charles River Breeding Laboratories (Wilmington, MA). In some experiments, we have used mice in which the B7-1 or the B7-2 gene was disrupted (B7-1 knockout (KO)5 or B7-2 KO mice, respectively) (19, 20) and which were bred back to BALB/c mice (that are H-2d like DBA/2 mice) for 10 generations. In these experiments, we have also used wild-type BALB/c mice (Charles Rivers Breeding Laboratories) as controls.

Tumor cells

All studies employed the B7-1 transfectant of P815 tumor cells (H-2d), which was previously described (13), that was maintained in vitro in low-glucose DMEM (Life Technologies, Grand Island, NY) supplemented with 10% FBS (Sigma, St. Louis, MO), 1% penicillin and streptomycin, and 0.8 mg/ml G418 sulfate (Life Technologies). Routinely, mice were inoculated s.c. with 3.0–5.0 x 106 B7-1+ P815 tumor cells. In experiments assessing the expression of the B7-2 molecule on tumor cells grown in vivo, tumor nodules of mice inoculated s.c. with B7-1+ P815 tumor cells were excised when the tumors reached 3–6 mm in diameter, and a single-cell suspension was prepared by treating tumor pieces with 1.4 mg/ml collagenase type I and 0.3 mg/ml DNase type I (Sigma) in DMEM, according to the method of Peters et al. (21) as modified by Berd et al. (22).

Flow cytometric analysis

Assessment of B7-1 and B7-2 expression on B7-1+ P815 tumor cells grown in vivo was conducted with FITC-conjugated anti-B7-1 mAb and phycoerythrin (PE)-conjugated anti-B7-2 mAb (both of which were purchased from PharMingen, San Diego, CA). The tumor cells were identified based on light scatter properties and intensity of B7-1 staining (i.e., the product of the transfected B7 gene). In experiments assessing up-regulation of B7-1 and B7-2 expression on host APCs, lymph node cells derived from three to five mice per group were stained simultaneously for B7 expression (with PE-conjugated anti-B7-1 or anti-B7-2 mAb) and for the expression of B220, MAC-1, or CD3 molecules (by the use of the appropriate FITC-conjugated mAb purchased from PharMingen). Finally, flow cytometric analysis of 20,000 viable cells was conducted on EPICS Elite ESP (Coulter, Hialeah, FL). Each experiment was repeated at least three times, and the results of a representative experiment are provided in the form of a histogram.

mAb treatments

In experiments assessing the importance of B7-1 and/or B7-2 expression for the rejection of B7-1+ P815 tumor cells, mice were given an i.p. injection of either: 1) anti-B7-1 (16-10A1 (23)), 2) anti-B7-2 (GL-1 (19)), 3) anti-B7-1 plus anti-B7-2, or 4) normal rat IgG (NIgG; Sigma) at a dosage of either 50 or 100 µg/mouse starting 2 h before tumor inoculation and every other day thereafter for the duration of the experiment. In experiments assessing the importance of CD4+ and/or CD8+ T cells for the rejection of B7-1+ P815 tumor cells, mice were given an i.p. injection of either 1) anti-CD4 (GK1.5), 2) anti-CD8 (116-13.1), 3) anti-CD4 plus anti-CD8, or 4) NIgG at a dosage of 1 mg/mouse starting 2 days before tumor inoculation and every 4–5 days thereafter for the duration of the experiment. This protocol of anti-CD4 and anti-CD8 mAb treatments was found (by indirect immunofluorescence staining followed by flow cytometric analysis of cells from the draining lymph nodes of the mice) to lead to >95% depletion of CD4+ cells or >90% depletion of CD8+ cells, respectively, with no decrease, but actually some increase, in the percentage of the other T cell subset. In all experiments, tumor growth was monitored three times a week with the aid of calipers, and the average diameter of two perpendicular measurements was determined. In adherence to animal care policy, mice were sacrificed when their tumor diameter approached 20 mm or the mice became morbid. Each experiment was repeated at least twice with 5–10 mice per group, and the results of a representative experiment are provided as mean tumor diameter for all mice in a given group ± SE.

Statistical analysis

To determine the significance of differences in the fraction of mice surviving following different treatments, the generalized Savage (Mantel-Cox) test was used. For all other statistical analyses, Student’s t test was employed. A p value of 0.05 or lower was considered significant in both tests.


   Results
Top
 Abstract
 Introduction
 Materials and Methods
 Results
Discussion
 References
 
Importance of B7-2 expression for the eradication of B7-1+ P815 tumor cells

Experiments were conducted to determine whether the ability of normal mice to reject P815 tumor cells transfected with the B7-1 gene depends only on the B7-1 molecule or also on the other member of the B7 family. Accordingly, we determined if treatment of mice with anti-B7-1 and/or anti-B7-2 mAb can slow down or even prevent the eradication of B7-1+ P815 tumor cells. For this purpose, the mice were treated with anti-B7-1 and/or anti-B7-2 mAb at a dosage of 100 µg/mouse every other day starting 2 h before tumor inoculation. As seen in Fig. 1Go, while anti-B7-1 mAb alone was effective in preventing the eradication of B7-1-transfected P815 tumor cells in all but one mouse, anti-B7-2 mAb alone did not prevent or slow down B7-1+ P815 tumor regression. However, anti-B7-2 mAb treatment facilitated tumor growth in mice treated with anti-B7-1 mAb, suggesting that B7-2 expression also contributes to the eradication of B7-1-transfected tumor cells.



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  		FIGURE 1. Effect of anti-B7-1 and/or anti-B7-2 mAb treatment on the mean tumor diameter of normal DBA/2 mice inoculated s.c. with B7-1+ P815 tumor cells. Mice were inoculated s.c. with 3 x 106 B7-1+ P815 tumor cells in conjunction with i.p. treatment with anti-B7-1 (•), anti-B7-2 ({blacktriangledown}), or anti-B7-1 plus anti-B7-2 ({blacktriangleup}) mAb at a dosage of 100 µg/mouse every other day for the duration of the experiment. Up to day 11 after tumor inoculation, all 10 mice treated with anti-B7-1 mAb were monitored as a single group. From day 14 on, the single mouse that showed clear tumor regression (1 of 10) was monitored separately from the rest of the mice that showed significant tumor progression (9 of 10). As a reference point, we provide information regarding the mean tumor diameter of mice inoculated s.c. with B7-1+ P815 tumor cells in conjunction with i.p. treatment with NIgG ({square}). The mean tumor diameter of mice receiving anti-B7-1 plus anti-B7-2 mAb was significantly larger (p <= 0.05) than that of mice receiving anti-B7-1 mAb alone from day 8 after tumor inoculation until the end of the observation period.

 
To better discern the importance of the B7-2 molecule for the eradication of B7-1+ P815 tumor cells, we used conditions under which the contribution of the B7-1 molecule was reduced (but not abrogated). Specifically, we utilized a less aggressive regimen of anti-B7-1 mAb treatment (which consisted of a lower dose of mAb, i.e., 50 µg instead of 100 µg, given every other day) that did not prevent the regression of B7-1+ P815 tumors, and determined whether administration of anti-B7-2 mAb (100 µg given every other day) to mice treated with the less aggressive regimen of anti-B7-1 mAb would be manifested in a profound delay in tumor regression and conceivably even in prevention of tumor regression. As seen in Fig. 2Go, treatment with the less aggressive regimen of anti-B7-1 or with anti-B7-2 mAb did not prevent or slow down B7-1+ P815 tumor regression, although the tumors reached a larger size before starting to regress in mice treated with anti-B7-1 mAb. In contrast, treatment of the mice with the combination of the less aggressive regimen of anti-B7-1 mAb plus anti-B7-2 mAb prevented tumor regression in all mice. Thus, not only the B7-1 molecule, but also the B7-2 molecule, contributes to the eradication of B7-1+ P815 tumor cells.



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  		FIGURE 2. Effect of a less aggressive regimen of anti-B7-1 mAb treatment and/or anti-B7-2 mAb treatment on the mean tumor diameter of normal DBA/2 mice inoculated s.c. with B7-1+ P815 tumor cells. Mice were inoculated s.c. with 3 x 106 B7-1+ P815 tumor cells in conjunction with i.p. treatment with 50 µg anti-B7-1 (•), 100 µg anti-B7-2 ({blacktriangledown}), or 50 µg anti-B7-1 plus 100 µg anti-B7-2 ({blacktriangleup}) mAb every other day for the duration of the experiment. As a reference point, we provide information regarding the mean tumor diameter of mice inoculated s.c. with B7-1+ P815 tumor cells in conjunction with i.p. treatment with NIgG ({square}). The mean tumor diameter of mice receiving anti-B7-1 plus anti-B7-2 mAb was significantly larger (p <= 0.05) than that of mice receiving anti-B7-1 or anti-B7-2 mAb alone from day 12 after tumor inoculation until the end of the observation period.

 
Lack of up-regulation of B7-2 expression on B7-1+ P815 tumor cells grown in vivo

In principle, the B7-2-expressing cell types that contributed to the eradication of B7-1+ P815 tumor cells by normal mice could have been host cells and/or B7-1+ P815 tumor cells that were induced in vivo to express the B7-2 molecule. Initially, we focused our attention on the tumor cells in light of reports that in some situations B7-negative tumor cells can be induced in vivo to express the B7-1 and the B7-2 molecules (24). Accordingly, tumor cells derived from the tumor nodules of mice inoculated s.c. with B7-1+ P815 tumor cells were subjected to two-color immunofluorescence staining followed by flow cytometric analysis. Specifically, tumor cells identified based on light scatter properties and bright staining for B7-1 transgene expression were examined for B7-2 expression. As seen in Fig. 3Go, the B7-1+ P815 tumor cells were negative for B7-2 expression, indicating that B7-1+ P815 tumor cells are not induced in vivo in the s.c. tumor nodules to express B7-2 molecules.



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  		FIGURE 3. Flow cytometric analysis of B7-2 expression on cells identified as B7-1+ P815 tumor cells among cells derived from the s.c. tumor nodules of B7-1+ P815 tumor-bearing mice. Tumor cells derived from the s.c. tumor nodules of DBA/2 mice inoculated with B7-1+ P815 tumor cells were stained with FITC-conjugated anti-B7-1 mAb and PE-conjugated anti-B7-2 mAb. The tumor cells were identified based on light scatter properties and intensity of staining for B7-1 expression. The data are presented as a single-color histogram for B7-2 expression on cells identified as B7-1+ P815 tumor cells (). As a reference point, we provide the histogram for B7-1+ tumor cells stained with PE-conjugated NIgG ().

 
Up-regulation of B7-2 expression on APCs from the draining lymph nodes of mice inoculated with B7-1+ P815 tumor cells

Studies were next undertaken to determine whether s.c. inoculation of B7-1+ P815 tumor cells into normal mice leads to up-regulation of B7-2 expression on host cells from the draining lymph nodes. Flow cytometric studies showed that B7-2 expression was up-regulated on cells from the draining lymph nodes on day 5 after B7-1+ P815 tumor inoculation (data not shown). Therefore, subsequent studies examining B7-2 expression on individual cell subsets (i.e., B220+, MAC-1+, or CD3+ cells) from the draining lymph nodes of mice inoculated with the B7-1+ P815 tumor cells were conducted on day 5 after tumor inoculation. As illustrated in Fig. 4Go, inoculation of B7-1+ P815 tumor cells into normal mice led to a substantial increase in the percentage of B220+ cells so that they represented approximately 35% of the cells among the draining lymph nodes as compared with <20% of the cells among lymph nodes from normal mice. This increase in the percentage of B220+ cells in the draining lymph nodes of mice inoculated with B7-1+ P815 tumor cells was associated with a decrease in the percentage of CD3+ cells so that they represented ~60% of the lymph node cells as compared with ~80% of the lymph node cells from normal mice. In addition, inoculation of B7-1+ P815 tumor cells led to a substantial increase in the percentage of B7-2+/B220+ cells among the draining lymph nodes (i.e., from 2.3% to 11.1%). This translates into an increase in the percentage of cells expressing the B7-2 molecule among the B220+ cells from ~13% to ~31%. At the same time, inoculation of B7-1+ P815 tumor cells was not associated with an increase in the percentage of B7-2+/MAC-1+ cells or B7-2+/CD3+ cells. Thus, inoculation of B7-1+ P815 tumor cells leads to up-regulation of B7-2 expression on host APCs, such as B220+ cells, from the draining lymph nodes.



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  		FIGURE 4. Two-color flow cytometric analysis of B7-2 expression on B220+ cells (A), MAC-1+ cells (B), and CD3+ cells (C) from the draining lymph nodes of mice inoculated s.c. with B7-1+ P815 tumor cells. Lymph node cells from normal mice (top histograms in each panel) or mice inoculated 5 days earlier with B7-1+ P815 tumor cells (bottom histograms in each panel) were stained with PE-conjugated anti-B7-2 mAb plus FITC-conjugated anti-B220 mAb, FITC-conjugated anti-MAC-1 mAb, or FITC-conjugated anti-CD3 mAb. As reference point, we provide the histograms for lymph node cells stained with PE-conjugated NIgG plus FITC-conjugated anti-B220 mAb, FITC-conjugated anti-MAC-1 mAb, or FITC-conjugated anti-CD3 mAb. Percentages of gated cells are shown in each histogram.

 
Up-regulation of B7-1 expression on APCs from the draining lymph nodes of mice inoculated with B7-1+ P815 tumor cells

Experiments were conducted to determine whether inoculation of B7-1+ P815 tumor cells into normal mice also leads to up-regulation of B7-1 expression on host cells from the draining lymph nodes. Flow cytometric studies showed that B7-1 expression was up-regulated on cells from the draining lymph nodes on day 7 after B7-1+ P815 tumor inoculation (data not shown). Therefore, subsequent studies examining B7-1 expression on individual cell subsets (i.e., B220+, MAC-1+, or CD3+ cells) from the draining lymph nodes of mice inoculated with the B7-1+ P815 tumor cells were conducted on day 7 after tumor inoculation. As seen in Fig. 5Go, on day 7 after B7-1+ P815 tumor inoculation, as on day 5 after B7-1+ P815 tumor inoculation (Fig. 4Go), the draining lymph nodes contained an increased percentage of B220+ cells and a decreased percentage of CD3+ cells relative to lymph nodes from normal mice. In addition, the draining lymph nodes from mice inoculated with B7-1+ P815 tumor cells contained a slightly higher percentage of B7-1+/B220+ cells as well as B7-1+/MAC-1+ cells, but not B7-1+/CD3+ cells, than lymph nodes from normal mice. However, while the increase in the percentage of B7-1+/B220+ cells does not translate into an increase in the percentage of cells expressing the B7-1 molecule among the B220+ cells, the increase in the percentage of cells expressing the B7-1 molecule among MAC-1+ cells translates into an increase in the percentage of cells expressing the B7-1 molecule among the MAC-1+ cells (i.e., from ~38% to ~48%). Thus, inoculation of B7-1+ P815 tumor cells leads to up-regulation of B7-1 expression on host APCs, such as MAC-1+ cells, from the draining lymph nodes.



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  		FIGURE 5. Two-color flow cytometric analysis of B7-1 expression on B220+ cells (A), MAC-1+ cells (B), and CD3+ cells (C) from the draining lymph nodes of mice inoculated s.c. with B7-1+ P815 tumor cells. Lymph node cells from normal mice (top histograms in each panel) or mice inoculated 7 days earlier with B7-1+ P815 tumor cells (bottom histograms in each panel) were stained with PE-conjugated anti-B7-1 mAb plus FITC-conjugated anti-B220 mAb, FITC-conjugated anti-MAC-1 mAb, or FITC-conjugated anti-CD3 mAb. As reference point, we provide the histograms for lymph node cells stained with PE-conjugated NIgG plus FITC-conjugated anti-B220 mAb, FITC-conjugated anti-MAC-1 mAb, or FITC-conjugated anti-CD3 mAb. Percentages of gated cells are shown in each histogram.

 
Importance of B7-expressing host APCs for the eradication of B7-1+ P815 tumor cells

To determine the importance of B7-1 and B7-2 expression on host APCs for the eradication of B7-1+ P815 tumor cells, we employed mice in which the B7-1 or B7-2 gene was disrupted (i.e., B7-1 KO or B7-2 KO mice, respectively) to prevent B7-1 or B7-2 expression on host APCs. Initially, we established that in B7-1 KO mice, like in wild-type DBA/2 mice, inoculation of B7-1+ P815 tumor cells leads to up-regulation of B7-2 expression on host APCs but not on tumor cells. Specifically, we established that B7-1+ P815 tumor cells derived from the s.c. tumor nodules of B7-1 KO mice do not express B7-2 molecules (Fig. 6Go). In addition, we established that inoculation of B7-1+ P815 tumor cells into B7-1 KO mice leads to an increase in the percentage of B7-2+/B220+ cells (but not B7-2+/CD3+ cells) in the draining lymph nodes (Fig. 7Go). Subsequently, we monitored B7-1+ P815 tumor growth/regression in B7-1 KO mice. As part of these studies, some of the B7-1 KO mice that were inoculated with B7-1+ P815 tumor cells were also treated with anti-B7-2 mAb to exclude the contribution of B7-2 expression on host APCs for B7-1+ P815 tumor eradication. As seen in Fig. 8Go, inoculation of B7-1+ P815 tumor cells into B7-1 KO mice, like inoculation of B7-1+ P815 tumor cells into wild-type mice, led to the appearance of tumors that grew transiently and then completely regressed. However, in contrast to our observations in control B7-1-expressing animals wherein the rate of B7-1+ P815 tumor regression was the same in wild-type mice treated with NIgG or with anti-B7-2 mAb, anti-B7-2 mAb treatment substantially delayed B7-1+ P815 tumor regression in B7-1 KO mice. Similar to our observations in the B7-1 KO mice, inoculation of B7-1+ P815 tumor cells into B7-2 KO mice also led to the appearance of tumors that grew transiently (Fig. 9Go). In addition, treatment of the B7-2 KO mice with a less aggressive regimen of anti-B7-1 mAb (i.e., 50 µg/mouse every other day) led to a substantial delay in the complete regression of B7-1+ P815 tumors relative to treatment with NIgG (Fig. 9Go). These results differ from our observations in the wild-type mice, wherein treatment with the less aggressive regimen of anti-B7-1 mAb did not lead to a delay in complete tumor regression relative to treatment with NIgG. Taken together, these results indicate that both B7-1- and B7-2-expressing host APCs are involved in the eradication of B7-1+ P815 tumor cells in wild-type mice.



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  		FIGURE 6. Flow cytometric analysis of B7-2 expression on cells identified as B7-1+ P815 tumor cells among cells derived from the s.c. tumor nodules of B7-1 KO mice. Tumor cells derived from the s.c. tumor nodules of B7-1KO mice inoculated with B7-1+ P815 tumor cells were stained with FITC-conjugated anti-B7-1 mAb and PE-conjugated anti-B7-2 mAb. The tumor cells were identified based on light scatter properties and intensity of staining for B7-1 expression. The data are presented as a single-color histogram for B7-2 expression on cells identified as B7-1+ P815 tumor cells (). As reference point, we provide the histogram for B7-1+ tumor cells stained with PE-conjugated NIgG ().

 


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  		FIGURE 7. Two-color flow cytometric analysis of B7-2 expression on B220+ cells (A) and CD3+ cells (B) from the draining lymph nodes of B7-1 KO mice inoculated s.c. with B7-1+ P815 tumor cells. Lymph node cells from naive B7-1 KO mice (top histograms in each panel) or B7-1 KO mice inoculated 5 days earlier with B7-1+ P815 tumor cells (bottom histograms in each panel) were stained with PE-conjugated anti-B7-2 mAb plus FITC-conjugated anti-B220 mAb or FITC-conjugated anti-CD3 mAb. As a reference point, we provide the histograms for lymph node cells stained with PE-conjugated NIgG plus FITC-conjugated anti-B220 mAb or FITC-conjugated anti-CD3 mAb. Percentages of gated cells are shown in each histogram.

 


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  		FIGURE 8. Comparison of B7-1+ P815 tumor regression following treatment with anti-B7-2 mAb vs NIgG in B7-1 KO mice of BALB/c background (H-2d) and in wild-type BALB/c or DBA/2 mice. Mice were inoculated s.c. with 5 x 106 B7-1+ P815 (H-2d) tumor cells in conjunction with i.p. treatment with anti-B7-2 mAb (•) or NIgG ({square}) at a dosage of 100 µg/mouse every other day for the duration of the experiment. The mean tumor diameter of mice receiving anti-B7-2 mAb was significantly larger (p <= 0.05) than that of mice receiving NIgG from day 4 until day 23 after tumor inoculation in the B7-1 KO mice, on day 12 after tumor inoculation in the wild-type DBA/2 mice, and at no time during the observation period in the wild-type BALB/c mice. As a reference point, we provide information regarding the mean tumor diameter of BALB/c mice inoculated with B7-1+ P815 tumor cells and treated with 100 µg/mouse of anti-B7-1 and 100 µg/mouse anti-B7-2 mAb every other day for the duration of the experiment ({blacktriangleup}). The mean tumor diameter of these mice was significantly larger (p <= 0.05) than that of BALB/c mice receiving NIgG or anti-B7-2 mAb from day 8 until day 27 after tumor inoculation.

 


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  		FIGURE 9. Comparison of B7-1+ P815 tumor regression following treatment with the less aggressive regimen of anti-B7-1 mAb vs NIgG in B7-2 KO mice of BALB/c background (H-2d) and in wild-type DBA/2 mice. Mice were inoculated s.c. with 5 x 106 B7-1+ P815 (H-2d) tumor cells in conjunction with i.p. treatment with anti-B7-1 mAb (•) or NIgG ({square}) at a dosage of 50 µg/mouse every other day for the duration of the experiment. The mean tumor diameter of mice receiving anti-B7-1 mAb was significantly larger (p <= 0.05) than that of mice receiving NIgG from day 7 until day 26 after tumor inoculation in the B7-2 KO mice, and from day 5 until day 12 after tumor inoculation in the wild-type DBA/2 mice.

 
Importance of CD8+ T cells, but not CD4+ T cells, for the eradication of B7-1+ P815 tumor cells

The previous results raised the question regarding the cellular basis of the tumor-eradicating immunity that is elicited by the B7-1+ P815 tumor cells and that depends on B7-expressing host APCs. Accordingly, we examined the possibility that the B7-expressing host APCs were presenting tumor-associated Ags to CD4+ T cells, which in turn promoted tumor eradication by CD8+ T cells. For this purpose, we determined whether anti-CD4 and/or anti-CD8 mAb treatment would prevent the rejection of B7-1+ P815 tumors by wild-type mice. As seen in Fig. 10Go, anti-CD8 mAb alone prevented tumor regression in all mice. In contrast, anti-CD4 mAb alone did not prevent or even slow down the regression of B7-1+ P815 tumors. Moreover, anti-CD4 mAb did not facilitate the rate of tumor progression in mice treated with anti-CD8 mAb. Thus, while CD8+ cells are essential for the eradication of B7-1+ P815 tumor cells, CD4+ T cells are apparently not involved.



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  		FIGURE 10. Effect of anti-CD4 and/or anti-CD8 mAb treatment on the mean tumor diameter of normal DBA/2 mice inoculated s.c. with B7-1+ P815 tumor cells. Mice were inoculated s.c. with 5 x 106 B7-1+ P815 tumor cells in conjunction with i.p. treatment with anti-CD4 ({blacktriangledown}), anti-CD8 (•), or anti-CD4 plus anti-CD8 ({blacktriangleup}) mAb at a dosage of 1 mg/mouse every 4–5 days for the duration of the experiment. As a reference point, we provide information regarding the mean tumor diameter of mice inoculated s.c. with B7-1+ P815 tumor cells and treated i.p. with NIgG ({square}). The mean tumor diameter of mice receiving anti-CD8 mAb alone or in conjunction with anti-CD4 mAb was significantly larger (p <= 0.05) than that of mice receiving either NIgG or anti-CD4 mAb alone from day 10 after tumor inoculation until the end of the observation period. The mean tumor diameter of mice receiving anti-CD8 mAb in conjunction with anti-CD4 mAb was not significantly larger (p >= 0.05) than that of mice receiving anti-CD8 mAb alone during the entire observation period.

 

   Discussion
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
References
 
Yang et al. (25) have recently shown that inoculation of B7-1+ P815 tumor cells into normal mice led to up-regulation of B7-2 expression on CD3- cells from the draining lymph nodes. Here, we confirm their observations with a different B7-1 transfectant of P815 tumor cells. In addition, we extend their observations by demonstrating that the CD3- cell type within the draining lymph nodes that displayed up-regulation of B7-2 expression was B220+. Moreover, we show that the B7-1-transfected P815 tumor cells also led to up-regulation of B7-1 expression on MAC-1+ cells in the draining lymph nodes of the mice. It is not known at present if the host APCs that displayed up-regulation of B7 expression following inoculation of B7-1+ P815 tumor cells were APCs that were present in the lymph nodes before tumor inoculation or represent APCs that migrated into the lymph nodes subsequent to the B7-1+ P815 tumor inoculation.

We show here that, although treatment of mice with the more aggressive regimen of anti-B7-1 mAb prevented the regression of B7-1+ P815 tumors, anti-B7-2 mAb treatment accelerated the rate of B7-1+ P815 tumor progression in the anti-B7-1-treated mice. These results indicate that not only B7-1 but also B7-2 molecules are involved in the eradication of B7-1+ P815 tumor cells. The importance of B7-2-expressing cells for the eradication of B7-1+ P815 tumor cells was even more apparent when a less aggressive protocol of anti-B7-1 mAb treatment was used, wherein anti-B7-1 mAb treatment alone was unable to prevent B7-1+ P815 tumor regression in any of the mice, yet anti-B7-2 mAb administration to such anti-B7-1-treated mice prevented tumor regression in all mice. The B7-2-expressing cells that contributed to the eradication of B7-1+ P815 tumor cells were most likely host APCs, since the B7-1+ P815 tumor cells were negative for B7-2 expression and were not induced in the s.c. tumor nodule to express the B7-2 molecule. At the same time, B7-1+ P815 tumor cells led to up-regulation of B7-2 expression on B220+ cells (but not CD3+ cells) from the draining lymph nodes. Moreover, the less aggressive protocol of anti-B7-1 mAb treatment that alone did not delay the complete B7-1+ P815 tumor regression in wild-type mice, delayed substantially the complete B7-1+ P815 tumor regression in mice that were unable to express the B7-2 molecule on host cells (B7-2 KO mice), indicating the importance of B7-2 expression on host cells for the eradication of B7-1+ P815 tumor cells.

B7-1 expression on host APCs is apparently also important for the eradication of B7-1+ P815 tumor cells. The importance of B7-1 expression on host APCs for the eradication of B7-1+ P815 tumor cells can be deduced from our studies in the B7-1 KO mice, as they allow us to distinguish between the contribution of B7-1 expression on tumor cells vs on host APCs. In these studies, anti-B7-2 mAb treatment, which on its own had no effect on the rate of B7-1+ P815 tumor regression in wild-type mice, substantially slowed down B7-1+ P815 tumor regression in the B7-1 KO mice, indicating the importance of B7-1 expression on host APCs (in addition to B7-2 expression on host APCs) for the eradication of B7-1+ P815 tumor cells.

Although up-regulation of B7 expression on host APCs is important for the acquisition of tumor-eradicating immunity against P815 tumor cells, a recent study by Yang et al. (25) illustrated that B7 expression on host APCs alone is not sufficient for the eradication of P815 tumors, and B7 expression on tumor cells is also required. Specifically, this study showed that although inoculation of B7-negative (parental) tumor cells led to up-regulation of B7 expression on host APCs in the draining lymph nodes, B7-negative P815 tumor cells, in contrast to B7-1+ P815 tumor cells, established progressively growing tumors (25). In addition, Yang et al. (25) concluded that B7 expression on host APCs is important for the generation of an antitumor immune response against B7-negative P815 tumor cells, as blockade of B7/CD28 interaction through the use of CTLA4Ig facilitated the growth of B7-negative P815 tumors. Given our current findings regarding the importance of B7 expression on host APCs for the eradication of B7-1+ P815 tumor cells, one of the functions of B7-1 expression on tumor cells is most likely to make tumor-associated Ags more readily available to host APCs for indirect Ag presentation. This may happen as a result of the ability of B7-1-expressing tumor cells to directly activate CD8+ T cells and NK cells (4, 9, 16, 26, 27), as well as a result of increased susceptibility of B7-1-expressing tumor cells to lysis by CTLs and NK cells (27, 28, 29).

Taken together, the above observations indicate that the tumor-eradicating immunity elicited by the B7-1+ P815 tumor cells is composed of three components. One component is the tumor-eradicating immunity elicited as a result of direct costimulation with B7-1 expressed on the P815 tumor cells. The second component is the tumor-eradicating immunity elicited as a result of costimulation with B7-1 expressed on host APCs, while the third component is the tumor-eradicating immunity elicited as a result of costimulation with B7-2 expressed on host APCs. The total tumor-eradicating immunity induced is apparently so powerful that when only B7-1 or B7-2 expression on host APCs is blocked, the tumor-eradicating immunity elicited by the B7-1 on the tumor cells plus the tumor-eradicating immunity elicited by the other member of the B7 family on host APCs is sufficient to cause complete tumor eradication at the regular rate. The contribution of B7-1 or B7-2 expression on host APCs is apparent when the contribution of both B7-1 and B7-2 expression on host APCs is blocked, as is the case in the anti-B7-2-treated B7-1 KO mice, or when the contribution of B7-1 expression on tumor cells and host APCs is reduced or eliminated by treatment with anti-B7-1 mAb.

Ectopic expression of B7-1 on tumor cells has been shown in several other tumor types to be effective in eliciting tumor-eradicating immunity (9, 10, 11, 12, 15). In most cases, CD8+ T cells, without help from CD4+ T cells, were sufficient for the eradication of B7-transfected tumor cells that expressed MHC class I, but not class II, molecules (10, 14, 16, 17, 18). These observations led to the conclusion that the B7-transfected tumor cells function as APCs and stimulate CD8+ T cells directly (4, 9). Here, we show that, in addition to the ability of B7-transfected tumor cells to stimulate CD8+ T cell-dependent tumor-eradicating immunity directly, such tumor cells can also stimulate CD8+ T cell-dependent tumor-eradicating immunity indirectly via cross-priming through B7-expressing host APCs. In this regard, it should be pointed out that although classically CD8+ T cells recognize Ags that are localized in the cytoplasm of target cells, processed, and presented as peptide complexes with MHC class I molecules, it is becoming increasingly apparent that exogenous Ags that are not expected to gain access to the cytoplasm can also be presented in association with MHC class I molecules (30, 31, 32). In this way, host APCs can stimulate CD8+ T cells against exogenous Ag.

The mechanisms through which the CD8+ T cells mediate their tumor-eradicating activity in vivo in mice inoculated with B7-transfected tumor cells remains to be elucidated. However, it is conceivable that CTL activity is one such mechanism, although CD8+ T cells may also produce cytokines that recruit and activate other effector cells, such as macrophages, to carry out tumor cell killing (33, 34, 35). In this regard, it is interesting to note that Huang et al. (36) have recently shown that, although B7-1+ colon carcinoma cells expressing influenza nucleoprotein can directly prime naive CTLs in vivo, most of the CTL priming in mice inoculated with these B7-1+ tumor cells is done by host APCs. It is not clear, however, whether CTL priming by host APCs occurred in their study without help from CD4+ cells, as is the case with the generation of tumor-eradicating immunity by CD8+ T cells in our studies.

Taken together, the results presented herein illustrate that B7-1-transfected tumor cells can stimulate the generation of tumor-eradicating immunity indirectly through presentation of tumor-associated Ags by B7-1- and/or B7-2-expressing host APCs, which in turn activate CD8+ T cells in the absence of help from CD4+ T cells. Consequently, our observations suggest that it may be possible to enhance the tumor-eradicating immunity elicited by B7-transfected tumor cells by combining this strategy with methods to enhance the differentiation and activation of host APCs (e.g., through the administration of granulocyte-macrophage CSF, IFN-{gamma}, and/or anti-IL-10 (31, 37)), thereby enhancing the effectiveness of B7-transfected tumor cells as tumor cell vaccines.


   Footnotes
 
1 This work is supported by research Grants P01 AI-35294, R01 AI-38310, and R01 CA-76532 from the National Institutes of Health. Back

2 In partial fulfillment of the requirements for the Doctor of Philosophy degree. Back

3 J.A.B. and M.B.M. are joint senior authors. Back

4 Address correspondence and reprint requests to Dr. Margalit B. Mokyr, Department of Biochemistry and Molecular Biology (M/C 536), University of Illinois, 1819 West Polk Street, Chicago, IL 60612. E-mail address: Back

5 Abbreviations used in this paper: KO, knockout; PE, phycoerythrin; NIgG, normal IgG. Back

Received for publication October 14, 1998. Accepted for publication January 26, 1999.


   References
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 References
 

  1. Janeway, C. A. J., K. Bottomly. 1994. Signals and signs for lymphocyte responses. Cell 76:275.[Medline]
  2. Jenkins, M. K., J. G. Johnson. 1993. Molecules involved in T-cell costimulation. Curr. Opin. Immunol. 5:361.[Medline]
  3. Bluestone, J. A.. 1995. New perspectives of CD28–B7-mediated T cell costimulation. Immunity 2:555.[Medline]
  4. Schwartz, R. H.. 1992. Costimulation of T lymphocytes: the role of CD28, CTLA-4, and B7/BB1 in interleukin-2 production and immunotherapy. Cell 71:1065.[Medline]
  5. Linsley, P. S., W. Brady, L. Grosmaire, A. Aruffo, N. K. Damle, J. A. Ledbetter. 1991. Binding of the B cell activation antigen B7 to CD28 costimulates T cell proliferation and interleukin 2 mRNA accumulation. J. Exp. Med. 173:721.[Abstract/Free Full Text]
  6. Harding, F. A., J. P. Allison. 1993. CD28–B7 interactions allow the induction of CD8+ cytotoxic T lymphocytes in the absence of exogenous help. J. Exp. Med. 177:1791.[Abstract/Free Full Text]
  7. Young, J. W., R. M. Steinman. 1990. Dendritic cells stimulate primary human cytolytic lymphocyte responses in the absence of CD4+ helper T cells. J. Exp. Med. 171:1315.[Abstract/Free Full Text]
  8. Chen, L., P. S. Linsley, K. E. Hellström. 1993. Costimulation of T cells for tumor immunity. Immunol. Today 14:483.[Medline]
  9. Allison, J. P., A. A. Hurwitz, D. R. Leach. 1995. Manipulation of costimulatory signals to enhance antitumor T-cell responses. Curr. Opin. Immunol. 7:682.[Medline]
  10. Townsend, S. E., J. P. Allison. 1993. Tumor rejection after direct costimulation of CD8+ T cells by B7-transfected melanoma cells. Science 259:368.[Abstract/Free Full Text]
  11. Chen, L., P. McGowan, S. Ashe, J. Johnston, Y. Li, I. Hellström, K. E. Hellström. 1994. Tumor immunogenicity determines the effect of B7 costimulation on T cell-mediated tumor immunity. J. Exp. Med. 179:523.[Abstract/Free Full Text]
  12. Chen, L., S. Ashe, W. A. Brady, I. Hellström, K. E. Hellström, J. A. Ledbetter, P. McGowan, P. S. Linsley. 1992. Costimulation of antitumor immunity by the B7 counterreceptor for the T lymphocyte molecules CD28 and CTLA-4. Cell 71:1093.[Medline]
  13. La Motte, R. N., M. A. Rubin, E. Barr, J. M. Leiden, J. A. Bluestone, M. B. Mokyr. 1996. Therapeutic effectiveness of the immunity elicited by P815 tumor cells engineered to express the B7-2 costimulatory molecule. Cancer Immunol. Immunother. 42:161.[Medline]
  14. Gajewski, T. F., F. Fallarino, C. Uyttenhove, T. Boon. 1996. Tumor rejection requires a CTLA4 ligand provided by the host or expressed on the tumor: superiority of B7-1 over B7-2 for active tumor immunization. J. Immunol. 156:2909.[Abstract]
  15. Matulonis, U., C. Dosiou, G. Freeman, C. Lamont, P. Mauch, L. M. Nadler, J. D. Griffin. 1996. B7-1 is superior to B7-2 costimulation in the induction and maintenance of T cell-mediated antileukemia immunity: further evidence that B7-1 and B7-2 are functionally distinct. J. Immunol. 156:1126.[Abstract]
  16. Wu, T. C., A. Y. Huang, E. M. Jaffee, H. I. Levitsky, D. M. Pardoll. 1995. A reassessment of the role of B7-1 expression in tumor rejection. J. Exp. Med. 182:1415.[Abstract/Free Full Text]
  17. Townsend, S. E., F. W. Su, J. M. Atherton, J. P. Allison. 1994. Specificity and longevity of antitumor immune responses induced by B7-transfected tumors. Cancer Res. 54:6477.[Abstract/Free Full Text]
  18. Johnston, J. V., A. R. Malacko, M. T. Mizuno, P. McGowan, I. Hellström, K. E. Hellström, H. Marquardt, L. Chen. 1996. B7-CD28 costimulation unveils the hierarchy of tumor epitopes recognized by major histocompatibility complex class I-restricted CD8+ cytolytic T lymphocytes. J. Exp. Med. 183:791.[Abstract/Free Full Text]
  19. Freeman, G. J., F. Borriello, R. J. Hodes, H. Reiser, K. S. Hathcock, G. Laszlo, A. J. McKnight, J. Kim, L. Du, D. B. Lombard, et al 1993. Uncovering of functional alternative CTLA-4 counter-receptor in B7-deficient mice. Science 262:907.[Abstract/Free Full Text]
  20. Sharpe, A. H.. 1995. Analysis of lymphocyte costimulation in vivo using transgenic and ’knockout’ mice. Curr. Opin. Immunol. 7:389.[Medline]
  21. Peters, L. C., J. S. Brandhorst, M. G. Hanna. 1979. Preparation of immunotherapeutic autologous tumor cell vaccines from solid tumors. Cancer Res. 39:1353.[Abstract/Free Full Text]
  22. Berd, D., H. C. Maguire, M. J. Mastrangelo. 1986. Induction of cell-mediated immunity to autologous melanoma cells and regression of metastases after treatment with melanoma cell vaccine preceded by cyclophosphamide. Cancer Res. 46:2572.[Abstract/Free Full Text]
  23. Razi-Wolf, Z., G. J. Freeman, F. Galvin, B. Benacerraf, L. Nadler, H. Reiser. 1992. Expression and function of the murine B7 antigen, the major costimulatory molecule expressed by peritoneal exudate cells. Proc. Natl. Acad. Sci. USA 89:4210.[Abstract/Free Full Text]
  24. Baskar, S., V. K. Clements, L. H. Glimcher, N. Nabavi, S. Ostrand-Rosenberg. 1996. Rejection of MHC class II-transfected tumor cells requires induction of tumor-encoded B7-1 and/or B7-2 costimulatory molecules. J. Immunol. 156:3821.[Abstract]
  25. Yang, G., M. T. Mizuno, K. E. Hellström, L. Chen. 1997. B7-negative versus B7-positive P815 tumor: differential requirements for priming of an antitumor immune response in lymph nodes. J. Immunol. 158:851.[Abstract]
  26. Nandi, D., J. A. Gross, J. P. Allison. 1994. CD28-mediated costimulation is necessary for optimal proliferation of murine NK cells. J. Immunol. 152:3361.[Abstract]
  27. Chambers, B. J., J. L. Wilson, M. Salcedo, K. Markovic, M. T. Bejarano, H. G. Ljunggren. 1998. Triggering of natural killer cell mediated cytotoxicity by costimulatory molecules. Curr. Top. Microbiol. Immunol. 230:53.[Medline]
  28. Geldhof, A. B., G. Raes, M. Bakkus, S. Devos, K. Thielemans, P. De Baetselier. 1995. Expression of B7-1 by highly metastatic mouse T lymphomas induces optimal natural killer cell-mediated cytotoxicity. Cancer Res. 55:2730.[Abstract/Free Full Text]
  29. Nishio, M., J. Spielman, R. K. Lee, D. L. Nelson, E. R. Podack. 1996. CD80 (B7.1) and CD54 (intracellular adhesion molecule-1) induce target cell susceptibility to promiscuous cytotoxic T cell lysis. J. Immunol. 157:4347.[Abstract]
  30. Rock, K. L.. 1996. A new foreign policy: MHC class I molecules monitor the outside world. Immunol. Today 17:131.[Medline]
  31. Brossart, P., M. J. Bevan. 1997. Presentation of exogenous protein antigens on major histocompatibility complex class I molecules by dendritic cells: pathway of presentation and regulation by cytokines. Blood 90:1594.[Abstract/Free Full Text]
  32. Albert, M. L., B. Sauter, N. Bhardwaj. 1998. Dendritic cells acquire antigen from apoptotic cells and induce class I-restricted CTLs. Nature 392:86.[Medline]
  33. Barth, R. J. J., J. J. Mule, P. J. Spiess, S. A. Rosenberg. 1991. Interferon {gamma} and tumor necrosis factor have a role in tumor regressions mediated by murine CD8+ tumor-infiltrating lymphocytes. J. Exp. Med. 173:647.[Abstract/Free Full Text]
  34. Fong, T. A., T. R. Mosmann. 1990. Alloreactive murine CD8+ T cell clones secrete the Th1 pattern of cytokines. J. Immunol. 144:1744.[Abstract]
  35. Zangemeister-Wittke, U., B. Kyewski, V. Schirrmacher. 1989. Recruitment and activation of tumor-specific immune T cells in situ: CD8+ cells predominate the secondary response in sponge matrices and exert both delayed-type hypersensitivity-like and cytotoxic T lymphocyte activity. J. Immunol. 143:379.[Abstract]
  36. Huang, A. Y., A. T. Bruce, D. M. Pardoll, H. I. Levitsky. 1996. Does B7-1 expression confer antigen-presenting cell capacity to tumors in vivo?. J. Exp. Med. 183:769.[Abstract/Free Full Text]
  37. Qin, Z., G. Noffz, M. Mohaupt, T. Blankenstein. 1997. Interleukin-10 prevents dendritic cell accumulation and vaccination with granulocyte-macrophage colony-stimulating factor gene-modified tumor cells. J. Immunol. 159:770.[Abstract]



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