Some characteristics of the cyclophosphamide-induced immunopotentiating cells in the spleen of mice bearing a large MOPC-315 tumor

We have shown previously (Ye, Q-W., and Mokyr, M. B. Cancer Res., 44: 3873-3879, 1984) that, following low-dose cyclophosphamide (CY) therapy (15 mg/kg) of mice bearing a large s.c. MOPC-315 tumor and extensive metastases, T-cell-dependent immunopotentiating activity appears in their hitherto immunosuppressive Sephadex G-10-adherent spleen cell population. Here we show that the CY-induced immunopotentiating T-cells express the Lyt 1, Lyt 2, and L3T4 phenotypes. The phenotype of the immunopotentiating T-cells was deduced from our observations that depletion of Lyt 1+, Lyt 2+, or L3T4+ cells from the Sephadex G-10-adherent spleen cell population of CY-treated tumor bearers abolished the ability of the adherent cells to enhance the generation of antitumor cytotoxicity when added to the in vitro immunization culture of normal spleen cells. Moreover, admixture of a Sephadex G-10-adherent cell population depleted of Lyt 2+ cells with a Sephadex G-10-adherent cell population depleted of L3T4+ cells failed to restore the immunopotentiating activity, indicating that T-cells that are apparently expressing simultaneously the Lyt 2 and L3T4 antigens are required for the exertion of the CY-induced immunopotentiating activity. The CY-induced immunopotentiating T-cells from MOPC-315 tumor bearers brought about the appearance of enhanced antitumor cytotoxicity not only against the MOPC-315 tumor cells, but also against two other syngeneic plasmacytomas, with surface immunoglobulin of a different class and antigenic specificity than the MOPC-315 tumor cells, as well as against a variant MOPC-315 tumor line which lacks surface immunoglobulin. The CY-induced immunopotentiating T-cells did not enhance the appearance of antitumor cytotoxicity against a syngeneic (WEHI 22.1) or an allogeneic (EL4) tumor of T-cell origin nor against the natural killer-sensitive YAC-1 cells. Thus, L3T4+, Lyt2+ T-cells from CY-treated MOPC-315 tumor bearers enhance the generation of antitumor cytotoxicity that is directed against plasmacytoma shared antigens other than immunoglobulins.     

Melphalan-induced enhancement of antitumor immune reactivity in thymocytes of adult BALB/c mice bearing a large MOPC-315 tumor

At no stage of tumor growth are thymocytes from MOPC-315 tumor bearers capable of bringing about the generation of enhanced antitumor cytotoxicity when added to immunization cultures of syngeneic normal spleen cells and "autochthonous" tumor cells. However, by Day 7 after low-dose melphalan [L-PAM (L-phenylalanine mustard)] therapy of mice bearing a large (greater than or equal to 20 mm) s.c. MOPC-315 tumor, their thymocytes exhibit such activity and it persists for at least 17 additional days. The ability of thymocytes from L-PAM-treated MOPC-315 tumor bearers to bring about the generation of enhanced antitumor cytotoxicity when added to immunization cultures of normal spleen cells and MOPC-315 tumor cells is evident over a 10-fold range of responder/stimulator cell ratios, and requires the presence of the thymocytes within the first day after initiation of the 5-day immunization cultures. In addition, immunization cultures containing normal spleen cells and thymocytes from L-PAM-treated MOPC-315 tumor bearers exhibit enhanced antitumor cytotoxicity by Day 4 after culture initiation that persists for at least 3 additional days. Thymocytes from L-PAM-treated MOPC-315 tumor bearers are able to bring about the generation of enhanced antitumor cytotoxicity only in response to stimulation with autochthonous tumor cells but not in response to stimulation with unrelated allogeneic EL4 tumor cells. The apparent specificity of the enhanced antitumor immune reactivity of thymocytes from L-PAM-treated MOPC-315 tumor bearers is not the result of extensive metastasis of tumor cells to the thymus. In fact, no tumor cells were found in the thymuses of MOPC-315 tumor bearers with methods that can detect 1 X 10(3) tumor cells, indicating that if MOPC-315 tumor cells metastasize at all into the thymus, the thymuses of mice bearing a large MOPC-315 tumor contain fewer than 1 X 10(3) tumor cells. Thus, thymocytes from mice which are engaged in the eradication of a large MOPC-315 tumor display enhanced antitumor immunity in response to stimulation with the autochthonous tumor cells. Such thymocytes may prove important to the outcome of low-dose L-PAM therapy for mice bearing a large MOPC-315 tumor, since the low-dose chemotherapy requires the contribution of T-cell-dependent antitumor immunity for its therapeutic effectiveness.     

Importance of Lyt 2+ T-cells in the curative effectiveness of a low dose of melphalan for mice bearing a large MOPC-315 tumor

We have previously demonstrated that the curative effectiveness of a low dose (2.5 mg/kg) of melphalan (L-phenylalanine mustard; L-PAM) for mice bearing a large s.c. (approximately 20 mm in diameter) MOPC-315 tumor and extensive metastases requires the participation of T-cell-dependent antitumor immunity in tumor eradication (S. Ben-Efraim et al., Cancer Immunol. Immunother., 15: 101-107, 1983). Here we show that the Lyt 2+ T-cells, and not the L3T4+ T-cells, participate in the cure of such tumor-bearing mice by a low dose of L-PAM. Specifically, depletion of Lyt 2+ T-cells from mice bearing a large MOPC-315 tumor by treatment with monoclonal anti-Lyt 2.2 antibody abolished the curative effectiveness of the low dose of drug. In contrast, depletion of L3T4+ T-cells from mice bearing a large MOPC-315 tumor by treatment with monoclonal anti-L3T4 antibody did not reduce significantly the curative effectiveness of the low dose of drug. Histological examination of tumor nodules on various days following low-dose L-PAM therapy revealed widespread lymphocytic infiltration by Day 5 following the chemotherapy, and this infiltration was drastically reduced when the L-PAM-treated tumor bearers were treated with either anti-Thy 1.2 or anti-Lyt 2.2 antibody but not with anti-L3T4 antibody. The antitumor immunity exhibited by Lyt 2+ T-cells derived from mice which were in the process of eradicating a large MOPC-315 tumor following low-dose L-PAM therapy was exploited successfully to confer systemic antitumor immunity to mice bearing a barely palpable tumor. Specifically, the adoptively transferred Lyt 2+ splenic T-cells, in conjunction with a subcurative dose of L-PAM, brought about the cure of most mice. The Lyt 2+ splenic T-cells from L-PAM-treated MOPC-315 tumor bearers were also found to be capable of exerting a direct potent lytic effect against MOPC-315 tumor cells in an antigen-specific manner. Thus, it is conceivable that the direct cytotoxic activity of Lyt 2+ T-cells for MOPC-315 tumor cells is responsible, at least in part, for the ability of the Lyt 2+ T-cells from L-PAM-treated MOPC-315 tumor bearers to bring about the eradication of the tumor burden not eradicated through the direct antitumor effects of the low dose of drug.     

Importance of Lyt-2+ T-cells in the resistance of melphalan-cured MOPC-315 tumor bearers to a challenge with MOPC-315 tumor cells

We have previously shown that mice bearing a large MOPC-315 tumor can be cured by a low dose of melphalan (L-phenylalanine mustard; L-PAM) if T-cell-dependent antitumor immunity also aids in tumor eradication (S. Ben-Efraim et al., Cancer Immunol. Immunother., 15: 101-107, 1983). Here we describe the phenotype of the T-cells that are responsible for the potent antitumor immunity exhibited by the L-PAM cured MOPC-315 tumor bearers. Initially, we identified the subset of T-cells responsible for the ability of spleen cells from L-PAM-cured MOPC-315 tumor bearers to neutralize tumor growth in the local adoptive transfer assay. The tumor-neutralizing activity of spleen cells from L-PAM-cured tumor bearers was drastically reduced when the spleen cells were depleted either in vitro or in vivo of Lyt-2+ cells. On the other hand, the tumor-neutralizing activity of spleen cells from L-PAM-cured MOPC-315 tumor bearers was not reduced, but actually enhanced, when the spleen cells were depleted either in vitro or in vivo of L3T4+ cells. The Lyt-2+ T-cells, and not the L3T4+ T-cells, were also found to be important for the ability of the intact L-PAM-cured MOPC-315 tumor bearers to reject a challenge with MOPC-315 tumor cells. Specifically, treatment of L-PAM-cured MOPC-315 tumor bearers with monoclonal anti-Lyt-2.2 antibody drastically reduced the ability of the mice to reject the tumor challenge. In contrast, treatment of the L-PAM-cured MOPC-315 tumor bearers with monoclonal anti-L3T4 antibody did not interfere with the ability of the mice to reject the tumor challenge, yet the same protocol of anti-L3T4 antibody treatment abolished the ability of splenic T-cells to provide help for the generation of a primary T-cell-dependent antibody response. The resistance of the L-PAM-cured MOPC-315 tumor bearers to challenge with MOPC-315 tumor cells was also dependent on the participation of carrageenan-sensitive effector cells, most likely macrophages, in tumor eradication. However, the immunity mediated by the T-cells, independent of carrageenan-sensitive effector cells, was extremely powerful and sufficient for the rejection of a tumor challenge with at least 300-fold, and possibly even 3000-fold, the minimal lethal dose of MOPC-315 tumor cells for 100% of normal mice.(ABSTRACT TRUNCATED AT 400 WORDS)     

Some approaches to improve the therapeutic effectiveness of adoptive chemoimmunotherapy with spleen cells from melphalan-treated BALB/c mice bearing a large MOPC-315 tumor.

Spleen cells from BALB/c mice that are in the process of eradicating a large MOPC-315 tumor following low-dose L-PAM therapy (L-PAM TuB spleen cells) were previously shown to be capable of bringing about the complete regression of a large (15 to 20 mm) s.c. MOPC-315 tumor in a substantial percentage of T-cell-deficient (athymic nude) mice that had been treated with low-dose L-PAM (adoptive chemo-immunotherapy; ACIT). Here we show that aggressive depletion of CD4+T-cells by treatment both of spleen-cell donors and of recipients with anti-L3T4 monoclonal antibody (MAb) greatly improved the therapeutic effectiveness of L-PAM TuB spleen cells in ACIT. In the light of the apparent importance of cytotoxic T-lymphocytes (CTLs) for tumor eradication in low-dose L-PAM-treated MOPC-315-tumor bearers, it is interesting that treatment of L-PAM TuB spleen-cell donors with anti-L3T4 MAb was found to result in the generation of enhanced splenic anti-MOPC-315 cytotoxicity. Although most athymic nude mice in which the tumor had apparently completely regressed following ACIT remained tumor-free, approximately 1/3 of the mice relapsed. However, a substantial percentage of the relapsing mice were rescued by a low dose of L-PAM, which was not effective in causing tumor regression in athymic nude mice bearing a comparably large tumor if the mice had not been subjected previously to ACIT. Almost all athymic nude mice that had been "cured" of a large MOPC-315 tumor by ACIT but did not resist a subsequent MOPC-315 tumor challenge were rescued by low dose L-PAM. Thus, the therapeutic effectiveness of L-PAM TuB spleen cells in ACIT may be improved by aggressive depletion of CD4+ T-cells, suggesting that a low dose of L-PAM, which leads to the acquisition of potent splenic-tumor-eradicating immunity in BALB/c mice bearing a large MOPC-315 tumor, does not eliminate completely (or possibly not at all) the inhibitory activity of CD4+ T-cells. In addition, athymic nude mice that are not endowed with fully protective tumor-eradicating immunity following ACIT still have a substantial residual anti-tumor immune potential that can be exploited to bring about eradication of a large tumor burden following low-dose L-PAM therapy.     

Importance of tumor-specific cytotoxic CD8+ T-cells in eradication of a large subcutaneous MOPC-315 tumor following low-dose melphalan therapy

We have previously demonstrated that depletion of CD8+ T-cells by the use of a monoclonal anti-Lyt-2.2 antibody abolishes the curative effectiveness of low-dose melphalan (L-phenylalanine mustard; L-PAM) therapy for BALB/c mice bearing a large (greater than or equal to 20 mm) s.c. MOPC-315 tumor and extensive metastases (Mokyr et al., Cancer Res., 49: 4597-4606, 1989). Here we show that as a consequence of low-dose L-PAM therapy, CD8+ T-cells accumulate in the s.c. tumor nodules of MOPC-315 tumor bearers. Specifically, an 80-fold increase in the number of CD8+ T-cells was seen within 5 days after the chemotherapy. Treatment of MOPC-315 tumor bearers with low-dose L-PAM in conjunction with monoclonal anti-Thy-1.2 or anti-Lyt-2.2 antibody, in contrast to treatment with monoclonal anti-L3T4 antibody, prevented the appearance of the massive CD8+ T-cell infiltrate in the s.c. tumor nodules. Fresh CD8+ T-cells derived from s.c. MOPC-315 tumor nodules that were regressing as a consequence of low-dose L-PAM therapy exhibited a potent direct lytic activity against the MOPC-315 plasmacytoma in a short-term in vitro assay. The specificity of the lytic activity exhibited by the CD8+ T-cells was illustrated not only by the inability of the CD8+ T-cells to lyse two antigenically unrelated thymomas (the WEHI 22.1 and the EL-4) and a natural killer-sensitive lymphoma (the YAC-1), but also by their relatively weak lytic activity against an antigenically related plasmacytoma (the MOPC-104E). Thus, CD8+ T-cells that infiltrate the s.c. tumor nodules of MOPC-315 tumor bearers following low-dose L-PAM therapy most likely exploit a CTL-type lytic mechanism to eradicate at least part of the large tumor burden not eliminated by the direct antitumor effects of the drug.     

Interleukin 2 requirement for the in vitro generation of antitumor cytotoxicity by thymocytes from melphalan-cured MOPC-315 tumor bearers

We have previously shown that enhanced antitumor cytotoxicity is generated when thymocytes from melphalan (L-phenylalanine mustard; L-PAM)-treated MOPC-315 tumor bearers, but not thymocytes from normal mice, are added to the immunization culture of syngeneic normal spleen cells and MOPC-315 tumor cells (Bartik et al., Cancer Res., 47: 4848-4855, 1987). Here we show that normal spleen cells produce, upon stimulation with MOPC-315 tumor cells, helper-like factors which are sufficient for thymocytes from L-PAM-treated MOPC-315 tumor bearers, but not for thymocytes from normal mice, to develop antitumor cytotoxicity in response to stimulation with MOPC-315 tumor cells. Since one of the helper-like factors produced by in vitro-immunized spleen cells is interleukin 2 (IL-2), we assessed the exogenous IL-2 requirements for the development of anti-MOPC-315 cytotoxicity in thymocytes from L-PAM-treated MOPC-315 tumor bearers, relative to thymocytes from normal mice. Thymocytes from L-PAM-treated MOPC-315 tumor bearers were found to require a 10-fold lower concentration of recombinant IL-2 (rIL-2) than thymocytes from normal mice in order to develop antitumor cytotoxicity in response to stimulation with MOPC-315 tumor cells. The concentration of rIL-2 required for the development of anti-MOPC-315 cytotoxicity by thymocytes from L-PAM-treated MOPC-315 tumor bearers was also 10-fold lower than the concentration of rIL-2 required by thymocytes from untreated MOPC-315 tumor bearers or thymocytes from L-PAM-treated normal mice. In addition, at any concentration of rIL-2 employed, thymocytes from L-PAM-treated MOPC-315 tumor bearers developed a higher level of anti-MOPC-315 cytotoxicity than did thymocytes from normal mice, L-PAM-treated normal mice, or untreated MOPC-315 tumor bearers. The enhanced antitumor cytotoxicity exhibited by thymocytes from L-PAM-treated MOPC-315 tumor bearers, following in vitro stimulation with MOPC-315 tumor cells plus rIL-2, was evident not only against MOPC-315 tumor cells but also against other syngeneic plasmacytomas but not an allogeneic thymoma. In addition, thymocytes from L-PAM-treated MOPC-315 tumor bearers required less rIL-2 than thymocytes from normal mice to develop antitumor cytotoxicity in response to stimulation with MOPC-315-associated antigens but not in response to stimulation with an allogeneic antigenically unrelated thymoma (EL4).(ABSTRACT TRUNCATED AT 400 WORDS)     

Production and enhanced anti-tumor activity of tumor necrosis factor in mice treated with cyclophosphamide

We demonstrated recently that the production of tumor necrosis factor (TNF) is induced in normal mice and in the immunosuppressed nude mouse model by the administration of muramyl dipeptide (MDP) derivatives followed by endotoxin (lipopolysaccharide). In the present study, the ability of this treatment to induce the production of TNF in mice receiving cyclophosphamide (CY) was examined. Two days following treatment with high-dose CY (250 mg/kg), mice exhibited leukocytopenia and drastically reduced splenic weight. However, these animals remained capable of producing TNF, albeit at lower levels, when treated with MDP derivatives and lipopolysaccharide (LPS), particularly when the lipophilic analogue MDP-dipalmitoyl glycerol (GDP) was utilized. TNF was also induced by the administration of MDP-GDP and LPS to Meth A sarcoma-bearing mice treated with this dose of CY. Furthermore, in all animals receiving this combination therapy, sarcoma necrosis and complete regression were obtained without any sign of tumor regrowth. A dose of 100 mg/kg CY was not effective for inhibiting tumor regrowth under the same experimental conditions. These results demonstrated that the anti-tumor activity of endogenously induced TNF is potentiated by combined therapy with a high dose of CY.     

人IL-12重组质粒对S-180荷瘤小鼠的治疗作用

目的：探讨人白细胞介素（imerleukin,IL）-12重组质粒对荷瘤小鼠的基因治疗作用．并进行初步的安全性评价。方法：建立肉瘤细胞（8-180）荷瘤小鼠模型,随机分为3组,每组10只,分别于注射瘤细胞后的第4、7、10、14和17天给予人IL-12重组质粒（pcDNA6-p70,100μg／只）、环磷酰胺（40mg／kg）及生理盐水（100μL／只）瘤体内注射。观察小鼠的存活时间;于致瘤后第21天处死小鼠,测肿瘤大小及质量,计算脾和胸腺指数;乳酸脱氢酶法检测NK杀伤活性;MTT法检测脾淋巴细胞增殖反应;ELISA法检测小鼠血清IFN-γ水平。对pcDNA6p70小鼠体内运用进行初步的安全性评价。结果：与生理盐水组相比．pcDNA6-p70瘤体内注射可显著延长荷瘤小鼠存活时间,P=0．03;缩小肿瘤体积,抑瘤率为30％,生理盐水组为0;增加脾（P=0．04）及胸腺（P=0．019）指数．增强其NK活性（P=0．001）及淋巴细胞增生反应（P=0．000）,提高荷瘤血清IFN-γ水平（P=0．000）。安全性检测结果显示,pcDNA6p70在小鼠体内应用无明显毒副作用．P=0．58。结论：人IL-12重组质粒可通过增强机体的细胞免疫功能发挥对肿瘤的基因治疗作用。     

Increase in alkaline phosphatase activity in the liver of mice bearing Ehrlich ascites tumor.

In mice bearing Ehrilich ascites tumors, alkaline phosphatase activity was increased fivefold in the liver and by 50% in the kidney. In mice bearing solid tumors caused by inoculation of tumor cells into the axillary region, the activity of this enzyme in the liver was increased 11-fold, whereas the activity in the kidney did not change. Alkaline phosphatase activities in the liver and kidney were not altered by administration of adrenal steroids. Adrenalectomy, fasting, and pregnancy did not affect the activity of alkaline phosphatase in the liver and kidney. Treatment with tumor extracts or ascites fluid of normal mice increased liver alkaline phosphatase activity. These findings suggested that the elevation of liver alkaline phosphatase activity was cuased primarily by the tumor itself, and not by hormonal imbalance provoked secondarily by the presence of the tumor.     

T-cell recruitment from the thymus to the spleen in tumor bearing mice: phenotypical alteration and recruitment of thymocytes raised in a tumor bearing state

Intrathymic events in mice bearing methylcholanthrene induced fibrosarcoma (MBS-1) were examined using mainly flow cytometric analysis. Ten days after tumor inoculation, the number of whole thymocytes was remarkably decreased. Surface phenotypical analysis by flow cytometry showed that the proportion of thymocytes with a low density of peanut agglutinin (PNAlow thymocytes), which is about 30% in normal mice, was increased to 70%. Histologically, the greater part of the thymus was occupied by the cortex. Moreover, the ratio of proliferating cells was increased in the PNAhigh cells. These findings in in vivo experiments suggested that a tumor bearing state would alter phenotypical characteristics of cortical thymocytes (PNAhigh, Thy 1high, H-2low) to medullary type (PNAlow, Thy 1low, H-2high). To support this hypothesis, in vitro experiments were performed using cultured thymic lymphoma EL4 cells, which possessed an immature thymus cell phenotype. Addition of serum from MBS-1 bearers to the culture medium of EL4 cells differentiated their phenotypical characteristics to the medullary type. Thus, it is assumed that factors in tumor bearers induce a massive migration of thymocytes by altering of phenotypes.     

Mode of action of polyethylene glycol 6000 in potentiating the in vitro generation of antitumor cytotoxicity by MOPC-315 tumor bearer spleen cells

Some of the possible mechanisms by which polyethylene glycol (PEG) augments the ability of MOPC-315 tumor bearer spleen cells to mediate in vitro antitumor cytotoxicity were evaluated. The level of antitumor cytotoxicity obtained in 5-day cultures of tumor bearer spleen cell suspensions correlated inversely with the percentage of Trinitrophenol (TNP)-rosettable cells (presumably metastatic tumor cells) present in the spleen. The kinetics of decrease in the percentage of TNP-rosettable cells coincided with the appearance of antitumor cytotoxicity. In addition, PEG was shown to interfere with the ability of viable tumor cells to suppress the in vitro generation of antitumor cytotoxicity in normal spleen cells cultured with mitomycin C-treated tumor cells. However, the decrease in the content of TNP-rosettable cells and the concurrent increase in the level of antitumor cytotoxicity were not due to direct cytotoxic and/or cytostatic effects of PEG on tumor cells. Spleen cells cultured in the presence of PEG had an increased rate of [3H]thymidine incorporation and proliferation compared to spleen cells cultured in the absence of PEG. However, the PEG-induced decrease in the percentage of TNP-rosettable cells either preceded or occurred at the same time that the PEG-induced increase in spleen cell number was observed. Therefore, spleen cell proliferation can at best explain only partially the PEG-induced decrease in the content of TNP-rosettable cells, and other mechanisms for the decrease must be considered.     

The effect of continuous infusion IL-1 alpha on carboplatin-induced thrombocytopenia and anti-tumor activity in RIF-1 tumor bearing mice

Interleukin-1 alpha (IL-1 alpha) has potent effects on hematopoiesis and can significantly enhance the anti-tumor activity of cytotoxic drugs. Studies were undertaken here to determine whether IL-1 alpha, administered by continuous infusion, could prevent carboplatin (CBDCA)-mediated thrombocytopenia and enhance CBDCA-mediated anti-tumor effects. RIF-1 tumor bearing mice were treated with CBDCA and IL-1 alpha either by a single bolus injection or by continuous infusion through the use of ALZET pumps. The duration and extent of CBDCA-induced thrombocytopenia in tumor-bearing mice was diminished when IL-1 alpha was administered continuously as compared to CBDCA alone or CBDCA plus a single bolus injection of IL-1 alpha. In addition, IL-1 alpha induced potentiation of CBDCA anti-tumor activity in vivo was significantly increased when IL-1 alpha was administered by continuous infusion. These results demonstrate the potential efficacy of IL-1 alpha by continuous infusion.     

Enhancement of antitumor activity of Propionibacterium avidum in combined with neurotropin in tumor bearing mice

The enhancement of antitumor activity of Propionibacterium avidum (P. avidum) in combination with Neurotropin (NSP) was investigated in C3H mice-MH 134 tumor system. P. avidum (0.5 mg) and NSP (20 mg/kg) were administered on day 2 and from day 1 to 7 after tumor inoculation, respectively. When mice were treated with P. avidum in combination with NSP, a significant prolongation in survival days was observed (P less than 0.01). Treatment with P. avidum alone produced prolongation in survival days, but NSP did fail. Increase of Con-A induced suppressor cell activity and depressed proliferative response of spleen cells were observed by the treatment with P. avidum. However, recovery of proliferative response to normal level and disappearance of suppressor cell activity were observed when NSP was combined. Thus, treatment by P. avidum in combination with NSP produced a significant prolongation in survival days and it may be depending on macrophage activation by P. avidum and on the restoration of T cell functions by NSP.     

The effect of tumor burden on ornithine decarboxylase activity in mice

Polyamines are essential for cell growth of normal and neoplastic tissue, alpha-Difluoromethylornithine (DFMO) is a known irreversible inhibitor or ornithine decarboxylase (ODC), the rate-limiting enzyme in polyamine biosynthesis. The purpose of this study was to examine the effects of tumor burden on ODC in tissues of tumor-bearing compared with tumor-free mice. Twenty-eight male Balb/c mice were divided into four groups of 7 each. Groups 1 and 2 were inoculated subcutaneously with 10 x 10(6) MC-26 mouse colon adenocarcinoma cells. Groups 3 and 4 were kept as tumor-free controls. Ten days after inoculation, groups 2 and 4 were injected with DFMO (200 mg/kg) intraperitoneally (IP) while Groups 1 and 3 received saline. Two hours after the injection of DFMO the animals were sacrificed. The tumor, pancreas, kidney, and liver were excised and analyzed for ODC activity. DFMO caused a significant reduction (compared with controls that did not receive DFMO) in the ODC activity of tumors; however, ODC activity of the kidney, pancreas, and liver of tumor-bearing mice was not affected. Additionally, the basal ODC activity in the kidney, liver, and pancreas of tumor-bearing mice was significantly lower compared with tumor-free controls. DFMO lowered ODC activity in the kidney, pancreas, and liver of tumor-free mice. These results suggest that the presence of MC-26 tumor causes systemic effects that alter ODC activity and the response to a known inhibitor of ODC.