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[Evaluation of the effectiveness of regional interleukin-2 dependent adoptive immunocytotherapy of mice with transplanted rhabdomyosarcoma].

Alternative principles of regional IL-2-dependent adoptive immunocytotherapy of malignant tumours are suggested. Regional adoptive immunocytotherapy consists in the administration of the short-term IL-2 treated lymphocytes of the lymph nodes removed from the tumour which have tumour-associated suppressor activity and spleen cells after elimination of cells with the immunosuppressive activity.

Animals↗

[Adoptive immunotherapy against peritoneal metastases from stomach cancer--application of regional lymph node lymphocytes].

Twenty-four patients with advanced stomach cancer were treated with OK-432 combined adoptive immunotherapy [AIT] as prophylaxis or therapy against peritoneal metastases. Lymphocytes isolated mainly from regional lymph nodes were cultured for 9-13 days with T cell growth factor and sonicated tumor extract. OK-432 administration and cell transfer were performed via a catheter inserted into the abdominal cavity at surgery. The proliferation rate of regional lymph node lymphocytes was higher than that of peripheral blood lymphocytes (p less than 0.01). Cultured regional lymph node lymphocytes expressed CD25 and CD4+45R- more frequently than those of peripheral blood lymphocytes. On the other hand, cytotoxic activity of regional lymph node lymphocytes were slightly lower than that of peripheral blood lymphocytes. These results suggest that regional lymph node lymphocytes could be used in AIT because of their different function from that of peripheral blood lymphocytes. Survival of the patients with peritoneal metastasis at primary laparotomy was higher than that of the historical control group (4-9 months after surgery). These results show that OK-432 combined adoptive immunotherapy appears to be a new therapeutic approach to peritoneal metastases from stomach cancer.

Adult↗

Adoptive immunotherapy of a rat glioma using lymphokine-activated killer cells and interleukin 2.

The aim of the present study was to develop an animal model to test the therapeutic potential of purified adherent lymphokine-activated killer (A-LAK) cells against an intracerebrally implanted rat glioma, designated F98. Highly purified A-LAK cells demonstrated greater activity against F98 tumor cells than conventional lymphokine-activated killer cells, as determined by means of 51Cr-release and clonogenic assays. Therapeutic efficacy was evaluated by means of a Winn neutralization assay, in which F98 targets and A-LAK cells or control nonadherent mononuclear cells were incubated for 18 h in vitro and then implanted stereotactically into the right caudate nuclei of Fischer rats. Animals given injections of 4000 F98 cells alone or control nonadherent mononuclear cells had a mean survival time of 22.3 days, compared to 46.1 days (P less than 0.001) for rats treated with A-LAK cells. Increasing the tumor inoculum to 12,500 cells reduced the survival time of A-LAK-treated animals to 27.8 days, compared to 20.8 days for untreated controls. Systemic administration of 50,000 units/kg of interleukin 2 every 12 h for 5 days failed to improve survival. The mean survival time of rats implanted with the F98 tumor ranged from 16 days for 10(5) cells to 29 days for 10(2) cells. Extrapolating from these survival data, treatment with A-LAK cells may have decreased the number of F98 cells to less than 10, but even this small number was still lethal. Supernatants from F98 cells had immunoinhibitory activity that, further, may have modulated the antitumor effects of A-LAK cells. Our results indicate that curative, adoptive immunotherapy of the F98 glioma by means of A-LAK/interleukin 2 is impossible to achieve and provide some explanation for the clinical failures that have been observed in the adoptive immunotherapy of malignant gliomas.

Animals↗

[Immunogenetic analysis of delayed hypersensitivity to tuberculin in mice by the method of local adoptive transfer].

It has been demonstrated that a specific phase of DTH reaction to tuberculin is carried out (and in adoptive transfer-transferred) by T-lymphocytes with Lyt-1+, Lyt-2-, L3T4+ phenotype by a method of local adoptive transfer of this reaction from BCG-immunized mice to the pads of intact recipients. From genetic viewpoint, the above reaction is limited by 1-A beta, 1-A alpha genes. In a syngeneic apposition, monoclonal antibodies anti-1-A kappa beta (without complement) were blocking the reaction transfer to CBA mice A beta chain affected mutation bm12 failed to limit the reaction transfer in apposition B6----B6.C-H-2bm12.

Animals↗

The immune response and the eye. III. Anterior chamber-associated immune deviation can be adoptively transferred by serum.

After the anterior chamber (AC) injection of trinitrophenol-coupled (TNP) spleen cells, it is observed that systemic delayed-type hypersensitivity responses to TNP are inhibited by Ag-specific suppressor T cells. We recently reported that suppression is initiated by viable TNP-coupled T cells within the inoculum and upon further analysis we found that these cells have the surface phenotype of CD4+ Ts inducer cells. We report here that treatment of these TNP-T cells with cycloheximide or cytochalasin-B before to AC injection abolishes suppression, whereas treatment with 2000 rad radiation does not. This indicates that protein synthesis and secretion are required to initiate suppression but proliferation is not. Further, we demonstrate the adoptive transfer of suppression by serum of AC inoculated animals. Detection of the component in serum in adoptive transfer assays, however, requires removal of the spleen before AC injection. We establish that the material in serum is a Ts cell product (T suppressor-inducer factor) based on three criteria: it is Ag specific, genetically restricted, and reactive with a mAb that specifically identifies these molecules. These results suggest that the signal leaving the eye to induce suppression of delayed-type hypersensitivity is T cell derived and that molecules mediating immune regulation for this organ are made within the eye and transported via the serum to the spleen.

Animals↗

In vivo expression of perforin by CD8+ lymphocytes in autoimmune disease. Studies on spontaneous and adoptively transferred diabetes in nonobese diabetic mice.

A potent cytolytic pore-forming protein (perforin or cytolysin) has previously been found to be associated with the cytoplasmic granules of CTL and NK cells. Inasmuch as all previous studies on perforin have been conducted with cultured CTL and NK cell lines, it is not clear whether perforin may play a role in the cytotoxicity mediated by CTL that have been primed in vivo. In this study, we investigated the presence of perforin in pancreata from nonobese diabetic (NOD) mice, which have been studied as a model of autoimmune, insulin-dependent (type I) diabetes mellitus. Whereas adult NOD mice spontaneously develop diabetes, it is possible to induce diabetes in young, irradiated NOD mice by adoptive transfer of splenocytes obtained from diabetic donors. By means of immunohistochemical analysis, we were able to detect perforin Ag in a small subpopulation of CD8+/Thy-1+/asialo GM1-/CD4- lymphocytes in the pancreatic islets of animals undergoing both spontaneous and adoptive transfer-mediated insulitis. Perforin+/CD8+ lymphocytes were found in small clusters and were observed to display the morphology of large granular lymphocytes. These observations show for the first time the presence of perforin-containing CD8+ lymphocytes in tissues of animals undergoing autoimmune disease.

Animals↗

Acquisition by the murine host of responsiveness toward various neoplastic cell lines, but not toward self, through adoptive transfer of a helper T-lymphocyte clone with antiself specificity.

Self-antigens, when expressed on neoplastic cells, have been shown to exhibit a certain antigenicity. We attempted to apply this antigenicity to enhance antitumor immune responses. Cells from the syngeneic, CD4+, CD8-, helper T-lymphocyte clone TE2 were adoptively transferred to C57BL/6 mice. TE2 lymphocytes recognize a self-antigen on splenocytes that is expressed aberrantly on the neoplastic cell lines EL4/8, EL4/13, B16-BL6, and PG19, all of C57BL/6 origin. Their adoptive transfer led to the rejection by the host of the former neoplastic cells and of 3LL carcinoma cells, administered 2 months later; inocula 40 to 80 times the minimal lethal size were rejected and conveyed to the mice a 10-fold enhanced cytotoxic T-lymphocyte response. Despite the autoimmune responsiveness of the TE2 T-lymphocytes, no graft-versus-host reaction was apparent. This conclusion is based on the absence of a polyclonal B-lymphocyte stimulation in the host, the stable number of residual donor TE2 cells, and the general health of the recipient mice. Consequently, the autoimmune and tumor-responsive TE2 cells, transplanted into the immune environment of the host, exhibit a specificity that is restricted toward neoplastic cells.

Animals↗

[Basic and clinical study of adoptive immunotherapy using cytotoxic T lymphocyte (CTL) against cancers].

Cytotoxic T cells (CTL) have been known to be one of the effector cells responsible for regression of tumors. In tumor-bearing hosts, CTL insufficiently attack the tumor because the suppressor cells inhibit the induction and activation of CTL. In our in vitro study, CTL were induced from peripheral blood lymphocytes (PBL) of cancer patients. These CTL showed the specific cytotoxic activity against autologous tumor in one case and broad specificity against tumors in other cases. A patient with breast cancer was treated with adoptive immunotherapy of CTL because she had severe side effect from antineoplastics . Her breast cancer was histologically scirrhous type adenocarcinoma which was resistant to antineoplastics. Patient's PBL were cocultured with mitomycin C treated-autologous tumor, and they were proliferated with interleukin 2 or T-cell growth factor (TCGF). Then, these CTL were injected to this patient intravenously at the interval of one or two weeks. Before serial injection of CTL, antineoplastics was prescribed in order to inhibit the function of suppressor cells and cancer cells. The sizes of tumors were gradually reduced, suggesting clinical regression. We suggest that combined treatment of adoptive immunotherapy using CTL and antineoplastics (CTL therapy) is very useful in the treatment of cancer patients.

Adenocarcinoma, Scirrhous↗

[An immunological and clinical evaluation of combined TAE-LAK adoptive immunotherapy].

To clarify the immunological effects of transcatheter arterial embolization (TAE) therapy for hepatocellular carcinoma (HCC), various immunological parameters were measured before and after TAE respectively. In most effective group of which AFP levels at first week after TAE therapy had decreased more than 50% compared with those before TAE, the percentage of OKT-4 and IL-2R positive cells in the peripheral blood lymphocytes (PBL) had significantly increased in number. In addition, IL-1, TNF, IL-2 and LAK activity were also enhanced by it. These immunological enhancement after TAE therapy was suggested to be a favourite condition for transferring LAK cells to the patients with HCC. Therefore the combination therapy with TAE and LAK-adoptive immunotherapy was conducted in 12 patients with HCC. Partial response to it was obtained in one case and minor response in three. However, no effectiveness was also found in eight (Progressive Disease: 1 case, No Change: 7 cases). Immunological response after combined TAE-LAK adoptive immunotherapy revealed that NK activity and LAK activity were markedly enhanced. Furthermore, the percentage of OKT-11+, IL-2R+ and Leu-7- 11c+ cells in the PBL had increased in number with statistically significant differences and OKT-8+ cells had increased in relative number. In conclusion, this study suggested that this combination therapy might be a well designed immunological and clinical therapy for HCC because it was done under the condition of well enhanced immunological parameters against tumors.

Carcinoma, Hepatocellular↗

Characterization of the antitumor activity of a polyantigenic immunomodulator (PAI): II--Involvement of NK cells and adoptive immunotherapy.

Natural killer (NK) cell activity and adoptive immunotherapy were used to characterize the anticancerous effect of a polyantigenic immunomodulator (PAI). PAI consists of a mixture of inactivated bacteria and influenza virus in a peanut oil-arlacel A-aluminum monoesterate emulsion, shown previously to have antitumoral activity in mice implanted with Ehrlich's ascites tumor. The administration of PAI, its bacterial or viral component strongly increased the in vitro activity of NK cells of splenocyte populations obtained from Swiss-Webster (allogeneic) and C57BL/6J (syngeneic) mice, specially during the early post-induction period. On the other hand, PAI-sensitized, allogeneic or syngeneic lymphocytes were transferred successfully to tumor-bearing mice implanted with Ehrlich's ascites tumor, reducing tumor growth and increasing survival. The results confirm our previous suggestions that PAI acts probably at the level of cellular immunity. Therefore complex polyantigenic substances such as PAI could be used directly alone, in combination with other immunoadjuvants or to sensitize in a global manner immunocompetent cells to be employed in adoptive immunotherapy.

Adjuvants, Immunologic↗

[Adoptive immunotherapy by intra-arterial infusion of ATLAK or Allo-TLAK cells in patients with head and neck cancer].

For clinical application of adoptive immunotherapy, it is necessary to prepare a sufficient number of autologous tumor specific effector cells. A large amount of peripheral blood lymphocytes was obtained by leukapheresis using a Heamonetics V50 pheresis system. Autologous tumor- and lymphokine-activated killer (ATLAK) cells were induced by autologous mixed lymphocyte tumor cell culture (autologous MLTC) and further activation with recombinant interleukin-2 (rIL-2). Another problem was the difficulty of obtaining a sufficient number of highly activated effector cells to reach the target tumor tissue. Direct infusion of effector cells into a feeding artery was effective for cell accumulation in the target. ATLAK cells were infused into maxillary artery in 4 patients with maxillary squamous cell carcinoma. The results indicated that the therapy was effective for reduction of the tumor mass. After immunotherapy, surgery was performed and the tissues were histologically examined. Degenerated tumor cells and intensive infiltration by mononuclear cells and macrophages were seen in the surrounding fibrous tissue. However, the quantity of fresh autologous tumor cells available from open biopsy was limited. Allogeneic cultured tumor cell line was used as stimulator of lymphocytes instead of autologous tumor cells. The killing activity of the allogeneic tumor and lymphokine activated killer (Allo-TLAK) cells was significantly induced against the autologous tumor cells. Antitumor effect was observed in 5 out of 9 patients. Side effects were minor, such as slight fever and blood eosinophilia, which may be due to the rIL-2 function. These results indicate that this method of therapy is an effective form of adoptive immunotherapy.

Adult↗

Adoptive immunity transferred by naive donor cells immunized in vitro.

Speedy restoration of immune responsiveness in bone marrow recipients has been the objective of studies in which the donor was immunized so that specific immunologic memory could be transferred adoptively and selectively. Using unrelated rabbits, matched for major histocompatibility antigens but mismatched for their immunoglobulin allotypes, it could be shown that recipients of lymphoid cells from naive donors became B cell chimeras but did not use donor-derived B cells for their antibody responses to test antigens. In contrast, cells from donors primed for such antigens dominated antibody production in recipients in response to specific challenge. Clonal restriction in such adoptive responses was demonstrated. We now show that the induction of effective memory in cells from naive donors can be achieved in vitro during the preparation of donor cells for transfer to the recipient. Early challenge of the recipient enhances expression of the transferred immune response quantitatively and also results in the establishment or preservation of a larger diversity of clones from the donor.

Animals↗

Adoptive chemoimmunotherapy of murine leukemia with helper T lymphocyte clones.

Antigen-specific syngeneic noncytolytic helper T lymphocyte clones were investigated for their ability to mediate successful adoptive chemoimmunotherapy (ACIT) of mice with established RBL5 tumors. Clone B10, specific for the viral coat m.w. 70,000 glycoprotein, could be rapidly activated in situ after local transfer with intact tumor cells in syngeneic hosts to produce a delayed-type hypersensitivity reaction. For ACIT, mice bearing 5-day-old tumors received cyclophosphamide followed by transfer of resting helper T lymphocyte clones with or without exogenous interleukin 2 (rIL-2). A single injection of clone B10 was effective in ACIT when followed by a short course of exogenous rIL-2. Alternatively, repeated injections of resting clone were also effective without exogenous rIL-2, suggesting that the major role for rIL-2 was prolongation of clone survival in vivo rather than activation of other effector cells. Clone F12, specific for a component of fetal calf serum, was not effective in ACIT either with or without rIL-2, even when administered under conditions known to result in clone activation. Thus, antigen-specific helper T lymphocyte clones are capable of activation and promotion of antitumor responses after adoptive transfer.

Animals↗

Adoptive immunity in immune-deficient scid/scid mice. I. Differential requirements of naive and primed lymphocytes for CD4+ T cells during rejection of minor histocompatibility antigen-disparate skin grafts.

Using a model system in which mature lymphocytes were adoptively transferred into immunodeficient C.B17-scid/scid recipients, the requirement for CD4+ T cells in rejection of previously healed-in multiple minor-H-antigen-disparate skin grafts was investigated. Depletion of functional CD4+ cells was accomplished by anti-CD4 antibody + complement treatment prior to adoptive transfer followed by chronic anti-CD4 serotherapy in vivo. Homozygous scid mice harboring nondepleted naive donor cells effectively rejected minor-H-antigen-disparate grafts, whereas scid/scid mice harboring CD4+ T-cell-depleted naive cells did not. Homozygous scid mice harboring minor-H-antigen-primed cells rejected the minor-H-antigen-disparate allografts regardless of whether or not the animals had received anti-CD4 treatment, although rejection by the mice receiving anti-CD4 treatment was retarded compared to mice not receiving anti-CD4 treatment. All the mice that received viable donor cells rejected minor + H-2 disparate allografts, demonstrating effective immunity in this case was not dependent upon the activity of CD4+ T cells. These data suggest that in responses against multiple minor-H-antigen-disparate tissue, primary allograft rejection is absolutely dependent upon CD4+ cells, and secondary allograft rejection is not. The implications of these findings in understanding interactions of T cell subsets in vivo and of the utility of using homozygous scid mice as recipients for exploring the function of transferred lymphoid cell populations are discussed.

Animals↗

Characterization of a cell population in thoracic duct lymph that adoptively transfers rejection of adult Trichinella spiralis to normal rats.

In Trichinella spiralis-infected rats, a population of cells in thoracic duct lymph (TDL) that can adoptively transfer protection to naive rats was identified and characterized. During the course of T. spiralis infection, blast cells appeared in lymph from Day 3-4, and only Day 3-4 TDL cells had protective properties after transfer. Protection was evident in a 1-2-day increase in the slow rejection of adult worms beginning 8-9 days after the challenge infection. The minimum number of TDL cells capable of transferring protection was 1.8 X 10(8) cells. Transferred cells could protect against a challenge infection with adult worms alone. A double cross-over experiment demonstrated that major histocompatibility complex identity was essential for effective transfer of protection (MHC restriction). An experiment using the mitotic inhibitor vinblastine showed that the protective cells belonged to a dividing cell population. The phenotype of the protective TDL was confirmed by a two-step cell separation procedure. First, it was demonstrated that surface Ig- cells (T cells) separated by affinity chromatography could transfer protection. Second, these surface Ig- cells were divided into two subpopulations by panning using monoclonal antibodies OX-8 and W3/25. The results showed that W3/25+ or OX-8- cells (T-helper) were effective in transferring protection. Protection was only seen when rats adoptively transferred with cells were challenged 1 day after cell transfer.

Animals↗

The chemistry and biology of unusual DNA structures adopted by oligopurine.oligopyrimidine sequences.

A family of unusual DNA structures has been discovered in segments with predominantly purines in one strand (pur.pyr sequences). These sequences are overrepresented in eukaryotic DNA and have been mapped near genes and recombination hot spots. When cloned into recombinant plasmids, many pur.pyr sequences are reactive to chemical and enzymic probes that are generally specific for single-stranded DNA. An intramolecular triplex is adopted by mirror repeats of G's and A's. Other non-B DNA structures adopted by similar sequences remain to be fully clarified but may be a family of related conformations. It is likely that these unorthodox structures play an important role in the function of the eukaryotic genome.

Animals↗

Fate of gamma-interferon-activated killer blood monocytes adoptively transferred into the abdominal cavity of patients with peritoneal carcinomatosis.

Five patients with colorectal cancer widely metastatic to peritoneal surfaces have been treated i.p. with infusions of autologous blood monocytes made cytotoxic by in vitro incubation with human gamma-interferon. The monocytes were purified by a combination of cytapheresis and counter-current centrifugal elutriation procedures; each week approximately 350 million activated monocytes were given to patients as adoptive immunotherapy by a single i.p. instillation. On the eighth cycle of treatment the trafficking of i.p. infused blood monocytes was studied in two patients by prelabeling the cells with 111In. These activated cells became distributed widely within the peritoneal cavity. Two and 5 days after infusion their position within the peritoneum had not changed. When peritoneal specimens were obtained 36 h after 111In-labeled monocyte infusion, labeled monocytes were demonstrated to be associated with the serosal surfaces by autoradiographic analysis. Scintiscanning structures outside the abdominal cavity revealed that 111In-labeled monocytes infused i.p. did not traffic to other organs during the 5 days of the study. We conclude that i.p. adoptive transfer of autologous killer blood monocytes is an effective way of delivering these cytotoxic cells to sites of tumor burden on peritoneal surfaces in these cancer patients.

Colonic Neoplasms↗

Immunity to a 3-methylcholanthrene-induced fibrosarcoma in the C57BL/6 mouse: in vivo analysis by the adoptive tumor neutralization test.

Analyses of the adoptive tumor neutralization test (modified Winn test) in C57BL/6 mice made immune to a 3-methylcholantheene-induced fibrosarcoma showed that the reaction was mediated by a thymus-derived lymphocyte, it was tumor-specific, and the resistance of the immunized donor mouse to the challenge was strongly correlated with the protection of the recipient mouse. Proliferation of immune cells and close contact between tumor cells and immune T-cells were required. The hypothesis of a participation of the recipient in the reaction was considered because of the lack of adoptive protection of pangenic nude mice.

Animals↗