Biomedical subjects

D Maccubbin

Publications and source records attributed to D Maccubbin.

8 recordsLinked to original sources

PubMed1995

Protective specific immunity induced by cyclophosphamide plus tumor necrosis factor alpha combination treatment of EL4-lymphoma-bearing C57BL/6 mice.

A combination treatment protocol initiated 12 days after tumor injection, when the tumor was large, by administering cyclophosphamide (CY, 150 or 250 mg/kg) intraperitoneally followed by intravenous tumor necrosis factor alpha (TNF alpha, 1000 units injection) on days 13, 16, 18, 21, and 23, resulted in about 60% long-term survival (i.e., survival for at least 60 days) in the syngeneic C57BL/6 mouse/EL4 lymphoma model system. The establishment of a specific antitumor immune memory and its possible therapeutic relevance was verified by reinjecting 60-day survivors with EL4 cells; all 60-day survivors that had received the combination treatments rejected the implants and survived for a further 60 days. Thymic cellularity was reduced during treatment and its recovery appeared to correlate with long-term survival and immunity. Thymocytes from mice treated with the combination were found to express significant levels of specific anti-EL4 cytolytic activity following a 4-day stimulation culture with X-irradiated EL4 cells and low concentrations of interleukin-2. This response could not be generated with thymocytes from naive animals. In each case the effect seen with the combination of a moderate CY dose (150 mg/kg) with TNF alpha was better than that seen with either dose of CY alone and equal to or better than that seen with the higher dose of CY combined with TNF alpha. These results indicate that treatment with a single moderate dose of CY in combination with TNF alpha is effective against a large, established tumor in this murine model. Furthermore, all the long-term survivors induced by this treatment developed protective immunity against reimplanted tumor and demonstrated a long-term specific immune memory in the thymus.

Adjuvants, Immunologic↗

PubMed1994

Intracellular doxorubicin kinetics in lymphoma cells and lymphocytes infiltrating the tumor area in vivo: a flow cytometric study.

Recently we have reported the development of a safe and effective chemoimmunotherapy protocol involving doxorubicin (Dox) in combination with interleukin 2 which, in C57BL/6 mice, boosts local T cell responses, and, in 50 to 80% of the cases, this resulted in the complete eradication of established syngeneic EL4 lymphoma or its Dox-resistant variant, EL4/A. Accumulation of host-derived leukocytes in the peritoneal cavity was increased up to 8-fold after tumor inoculation, but, in absolute numbers, did not increase further following Dox administration. The cellular pharmacokinetic studies undertaken to clarify the role of Dox following a single IV injection indicate that 4 h later, lymphocytes found in the peritoneal cavity have detectable levels of Dox; but the lymphoma cells (both EL4 and EL4/A) have, in proportion to their larger size, taken up more drug as judged by flow cytometry. The estimated drug "concentration" (i.e., intracellular amount divided by estimated cell size) at the 4-h time point, however, was found to be essentially equivalent in both the lymphoma cells and the lymphocytes. Thereafter, the drug content and intracellular "concentration" in the EL4/A cells rapidly declined while their numbers progressively increased. In contrast, the EL4 lymphoma cells and the lymphocytes found in the peritoneal cavity in the presence of either lymphoma consistently exhibited higher levels of drug 24-48 h than at 4 h. Splenic and tumor-infiltrating mature T (CD3+) cells were completely insensitive to Dox cytotoxicity and actually showed increased CTL activity when examined ex vivo. Although EL4 cells had identical Dox uptake patterns to those of CD3+ cells, they were sensitive to the drug and their numbers decreased, resulting in increased host/tumor cell ratios in these mice. The pharmacokinetic parameters of the drug and the insensitivity of the mature T cells to the drug determined in this study can explain, in part, the efficacy of a chemoimmunotherapy protocol boosting local T-cell responses.

Animals↗

PubMed1994

TNF-alpha potentiation of the lymphokine-activated killer response of murine thymus cells.

The role of tumor necrosis factor (TNF-alpha) on in vitro generation of lymphokine-activated killer (LAK) cells from murine thymocytes was investigated and compared to that on generation of LAK from splenocytes. TNF-alpha increased the potential of interleukin-2 (IL-2) at suboptimal concentrations to generate LAK activity in thymocytes even more than in splenocytes. In parallel, augmented [3H]thymidine uptake by thymocytes and splenocytes was seen. However, no net increase in viable cell number was observed. LAK effector cells from TNF-alpha plus IL-2 cultures responded with an increased [3H]thymidine uptake to restimulation by IL-2 alone. These results suggest that TNF-alpha + IL-2 may be inducing the expansion of a small subset of cells. NK1.1+ cells are a very minor subset of thymocytes, nevertheless phenotype analysis showed that in thymocytes, IL-2 + TNF-alpha generates NK1.1+ CD8- LAK effectors in contrast to NK1.1- CD8+ cells found with IL-2 alone. This result is consistent with the finding in the proliferation studies. The fact that thymocytes are stimulated by the TNF-alpha + IL-2 combination to proliferate as well as to develop a phenotypically distinct effector supports the role of TNF-alpha in intra- and extrathymic regulation.

Animals↗

PubMed1993

Development of a safe and effective adriamycin plus interleukin 2 therapy against both adriamycin-sensitive and -resistant lymphomas.

This laboratory has extensively studied Adriamycin (doxorubicin)-induced immunomodulation. Despite demonstration of favorable effects, little therapeutic advantage was seen, and it was decided to test Adriamycin in combination with interleukin 2 (IL2). Considerable toxicity was seen with either high-dose IL2 or high-dose Adriamycin alone, using the syngeneic C57B1/6-EL4 T cell lymphoma model. When the doses of either agent were reduced to decrease toxicity, little therapeutic effect was seen. In contrast, an effective protocol without apparent toxicity was developed by combining a moderate dose of Adriamycin (4 mg/kg, IV, Days 1 and 8 or only Day 8) with prolonged administration of a moderate dose of IL2 (2 micrograms, b.i.d., i.p., Days 9 to 40). This protocol resulted in up to 80% long-term survivors among mice inoculated on Day 0 with EL4 lymphoma (5 x 10(4) cells). It should be noted that under these conditions, neither agent, when administered singly, induced long term survivors, and that following the inoculation of only 10-100 EL4 tumor cells all animals died in the absence of treatment. The survivors developed protective immunity as demonstrated by their ability to resist reimplantation with EL4 tumor. Furthermore, this resistance to tumor reimplantation could be transferred into naive hosts with spleen cells from tumor-bearing mice receiving the combination protocol; exposure of mice to sublethal whole body irradiation prior to tumor implantation completely abrogated the efficacy of this combination treatment. Finally, it was shown that this combination protocol was equally effective against an Adriamycin-resistant subline of EL4 that expresses the multidrug resistance phenotype.

Animals↗

PubMed1992

Tumoricidal activation of murine resident peritoneal macrophages by interleukin 2 and tumor necrosis factor alpha.

The capacity of recombinant human interleukin 2 (rH-IL2), alone or in combination with recombinant tumor necrosis factor (r-TNF alpha), to activate murine resident peritoneal macrophages to a tumoricidal state was examined. Resident peritoneal exudate cells from C57BL/6 mice were cultured for 18 h with activating agents and washed and the adherent cells (macrophages) were assessed for cytolytic activity against radiolabeled target tumor cells (EL4, P815). Under these conditions, rH-IL2 alone activated macrophages to a tumoricidal state in a concentration dependent fashion. Neither murine nor human r-TNF alpha alone had any activating effect but, when combined with rH-IL2, further stimulated rH-IL2-inducible responses. Using polymyxin B, it was shown that macrophage activation was not due to an inadvertent lipopolysaccharide contamination of the r-TNF alpha or rH-IL2 preparations. It was also unlikely that target cell lysis was a direct result of increased TNF alpha production by rH-IL2 stimulated macrophages since P815 is totally resistant to lysis by r-TNF alpha. Although the lytic effector function was mediated by adherent cells, nonadherent peritoneal exudate cells were required for activation to occur. Furthermore, antisera against murine gamma-interferon, when added to activation cultures, reduced the level of cytolytic activity which developed. These data suggest that rH-IL2-induced peritoneal macrophage activation requires stimulation of nonadherent cells and is dependent upon gamma-interferon mediated mechanisms.

Animals↗

PubMed1987

Effect of recombinant human tumor necrosis factor on the induction of murine macrophage tumoricidal activity.

The ability of recombinant human tumor necrosis factor (rH-TNF) alone or in combination with lymphokines (LK) to induce the in vitro activation of murine macrophages was evaluated. The treatment of C57BL/6 mouse resident peritoneal exudate cells (PEC) with rH-TNF and LK was found to induce the activation of macrophages to a tumoricidal state against P815 mastocytoma cells. Neither rH-TNF nor LK alone induced macrophage cytotoxic activity. Furthermore, the macrophage activation seen was not due to small amounts of contaminating lipopolysaccharide. The TNF plus LK-mediated macrophage activation could be totally ablated by rabbit antiserum to murine gamma-interferon, thus suggesting a role for gamma-interferon in this system. Since adherent cells (greater than or equal to 95% macrophages) only marginally responded to stimulation with rH-TNF plus LK and the addition of nonadherent PEC caused a marked augmentation of rH-TNF plus LK-mediated macrophage activation, the involvement of nonadherent PEC was suggested. In addition, using antibodies and complement to deplete subsets of cells from the nonadherent PEC, the requirement for cells bearing Thy 1.2 and asialo GM1 surface markers was demonstrated. These results suggest that TNF may play an autocrine regulatory role in concert with lymphokines in macrophage-mediated host defense against malignant neoplasia.

Animals↗

PubMed1987

Low concentrations of thymidine modulate allogeneic responses.

The effect of the addition of low concentrations of thymidine on the development of primary allogeneic immune responses in culture was studied. It was found that, dependent upon day of addition to culture, allogeneic cytotoxic T-lymphocyte generation was augmented by thymidine in the concentration range of 10(-9) to 10(-8) M while specific T-suppressor cell development was inhibited by 10(-7) M thymidine. Based on the concentration of thymidine in serum, its addition in the concentration range of 10(-9) to 10(-7) M results in 0.1 to 10 fold concentration change in cultures supplemented with 10% serum. While the mechanism(s) by which such minor changes in thymidine concentrations induced the modulations seen have not been elucidated, it is clear that such changes are easily attainable. In fact, such variations in the concentration of thymidine could contribute to the variable results often observed with different lots of fetal calf serum and workers investigating immunomodulations in culture systems should be aware of this possibility.

Animals↗

PubMed1986

Correlation between adriamycin-induced augmentation of interleukin 2 production and of cell-mediated cytotoxicity in mice.

Based on the observation that spleen cells from Adriamycin-treated mice could develop augmented levels of cytotoxic T-lymphocyte activity in response to heat-treated and/or X-irradiated alloantigens, it was postulated that modulations in soluble mediators could be involved in this phenomenon. In fact, in this study Adriamycin-induced increases in the levels of prostaglandin E2 and interleukin 2 activity have been observed with isolated cells. The "interleukin 2-like" activity was indistinguishable from that of partially purified interleukin 2 in terms of ability to restore responsiveness to experimentally inhibited primary alloantigen response cultures and to maintain long-term cultures of activated T-cells. Furthermore this latter activity was completely ablated by antiinterleukin 2 monoclonal antibody. While the modification in prostaglandin E2 production did not appear to play a role in determining augmentation of cytotoxic T-cell activity, the modification in interleukin 2 production was consistent with the possibility that this is a primary mechanism of Adriamycin-induced augmented cell-mediated cytotoxicity.

Animals↗