Evaluation of basic procedures for adoptive immunotherapy for gastric cancer

Peripheral blood lymphocytes (PBL) of gastric cancer patients in advanced stages showed lymphokine activated killer (LAK) activities comparable to those of healthy donors, suggesting potential applicability of LAK cells induced from PBL stimulated with recombinant interleukin-2 (rIL-2) in adoptive immunotherapy (AIT) for gastric cancer. In order to generate a large number of LAK cells from PBL, lymphocytes were cultured with both rIL-2 and phytohemagglutinin (PHA). In this culture, the numbers of cells increased to a greater extent than those in culture with rIL-2 alone but cytotoxic activity did not augment, thus suggesting that this procedure would not afford sufficient clinical effects. On the other hand, a large number of LAK cells with high anti-tumor activities were efficiently induced from spleen cells of the patients by culture of rIL-2; hence clinical usefulness of these LAK cells is anticipated. In regional lymph node lymphocytes (RLNL) cultured with rIL-2, the cytotoxic activities were lower than in those induced in PBL, and a characteristic increase of CD8 + CD11 + suppressor T cells was observed after incubation with rIL-2. Nevertheless, an increase of CD4 + 4B4⁺ helper inducer T cells was also observed in RLNL after the culture with rIL-2. Furthermore, high cytotoxic activities were induced in RLNL in some cases in which metastasis to the regional lymph nodes was not detected. When gastric cancer patients were pretreated with biological response modifiers (BRM), especially with Lentinan, LAK cells from PBL showed higher NK and LAK activities as compared with those of patients without BRM pretreatment.This work was partly supported by a grant from Hokkoku Cancer Research Foundation.     

Successful adoptive immunotherapy with vaccine-sensitized T cells,despite no effect with vaccination alone in a weakly immunogenic tumor model

Tumor cell vaccines have been successful at inducing immunity in naïve mice, but only in a few reports has vaccination alone induced regression of established tumors and, generally, only when they are very small. Clinically, vaccinations alone may not be able to cause regression of established human cancers, which tend to be weakly immunogenic. We hypothesized that pharmacologic ex vivo amplification of a vaccination-induced immune response with subsequent adoptive immunotherapy (AIT) to tumor-bearing animals would be more effective in treatment of these animals than vaccination alone. The 4T1 and 4T07 mammary carcinomas are derived from the same parental cell line, but 4T1 is much less immunogenic and more aggressive than 4T07. Vaccination with either 4T1, 4T1-IL-2, or 4T07-IL-2 was not effective as treatment for established 4T1 tumors. However, 4T1 or 4T07-IL-2-vaccine-sensitized draining lymph node (DLN) cells, activated ex vivo with bryostatin 1 and ionomycin and expanded in culture, induced complete tumor regressions when adoptively transferred to 4T1 tumor-bearing animals. This was effective against small tumors as well as more advanced tumors, 10 days after tumor cell inoculation. Furthermore, as would be required for this approach to be used clinically, vaccine-DLN cells obtained from mice with established progressive 4T1 tumors (inoculated 10 days before vaccination) also induced regression of 4T1 tumors in an adoptive host. In none of these experiments was exogenous IL-2 required to induce tumor regression. The response to tumor cell vaccine can be amplified by ex vivo pharmacologic activation of sensitized T cells, which can then cure an established, weakly immunogenic and highly aggressive tumor that was resistant to vaccination alone.     

Ox40 costimulation enhances the development of T cell responses induced by dendritic cells in vivo.

Dendritic cells (DCs) are bone marrow-derived APCs that display unique properties aimed at stimulating naive T cells. Several members of the TNF/TNFR families have been implicated in T cell functions. In this study, we examined the role that Ox40 costimulation might play on the ability of DCs to regulate CD4(+) and CD8(+) T cell responses in vivo. Administration of anti-mouse Ox40 mAb enhanced the Th response induced by immunization with Ag-pulsed DCs, and introduced a bias toward a Th1 immune response. However, anti-Ox40 treatment enhanced the production of Th2 cytokines in IFN-gamma(-/-) mice after immunization with Ag-pulsed DCs, suggesting that the production of IFN-gamma during the immune response could interfere with the development of Th2 lymphocytes induced by DCs. Coadministration of anti-Ox40 with DCs during Ag rechallenge enhanced both Th1 and Th2 responses induced during a primary immunization with DCs, and did not reverse an existing Th2 response. This suggests that Ox40 costimulation amplifies an ongoing immune response, regardless of Th differentiation potential. In an OVA-TCR class II-restricted adoptive transfer system, anti-Ox40 treatment greatly enhanced the level of cytokine secretion per Ag-specific CD4(+) T cell induced by immunization with DCs. In an OVA-TCR class I-restricted adoptive transfer system, administration of anti-Ox40 strongly enhanced expansion, IFN-gamma secretion, and cytotoxic activity of Ag-specific CD8(+) T cells induced by immunization with DCs. Thus, by enhancing immune responses induced by DCs in vivo, the Ox40 pathway might be a target for immune intervention in therapeutic settings that use DCs as Ag-delivery vehicles.     

C-Med 100®, a hot water extract of Uncaria tomentosa, prolongs lymphocyte survival in vivo

Water extracts of the bark of Uncaria tomentosa, a vine indigenous to South America, has been used for generations as an "immuno modulator". To understand the basis of this immuno modulatory effect we fed mice in their drinking water with C-Med 100, which is a commercially available water extract from Uncaria tomentosa. We found a dose-dependent increase in spleen cell numbers in the supplemented mice, but the proportions of B cells, T cells, NK cells, granulocytes, and memory lymphocytes were normal. However, there were no detectable changes of the lymphoid architecture of the spleen even after long-term treatment. Further, when C-Med 100 treatment was interrupted the cellularity returned to normal level within four weeks. The increased number of lymphocytes was most likely not due to increased production because C-Med 100 did not have any significant effect on precursor cells nor on the accumulation of recent thymic emigrants in the spleen. We conclude that accumulation is most likely due to prolonged cell survival, because adoptive transfer experiments demonstrated that C-Med 100 treatment significantly prolonged lymphocyte survival in peripheral lymphoid organs, without increasing their proliferation rate. Since the accumulation was reversible and without detectable pathological effects, these results suggest the use of C-Med 100 as a potential agent for clinically accelerating the recovery of patients from leukopenia.     

Augmentation versus inhibition: effects of conjunctional OX-40 receptor monoclonal antibody and IL-2 treatment on adoptive immunotherapy of advanced tumor.

Therapeutic efficacy of adoptive immunotherapy of malignancies is proportional to the number of effector T cells transferred. Traditionally, exogenous IL-2 treatment has been used to promote the survival and function of transferred cells. Recently, we described the therapeutic effects of in vivo ligation of the costimulatory receptor, OX-40R, on activated T cells during early tumor growth. In this study, we examined the effects of IL-2 and OX-40R mAb on adoptive immunotherapy of advanced tumors. For treatment of 10-day 3-methylcholanthrene 205 pulmonary metastases, systemic transfer of 50 x 10(6) activated tumor-draining lymph node T cells resulted in >99% reduction of metastatic nodules. With either IL-2 or OX-40R mAb conjunctional treatment, only 20 x 10(6) cells were required. Advanced 10-day 3-methylcholanthrene 205 intracranial tumors could be cured by the transfer of 15 x 10(6) L-selectin(low) T cells derived from draining lymph nodes. In this situation, IL-2 administration inhibited therapeutic effects of the transferred cells. By contrast, 5 x 10(6) T cells were sufficient to cure all mice if OX-40R mAb was administrated. Studies on trafficking of systemically transferred T cells revealed that IL-2, but not OX-40R mAb, impeded tumor infiltration by T cells. Tumor regression required participation of both CD4 and CD8 T cells. Because only CD4 T cells expressed OX-40R at cell transfer, direct CD4 T cell activation is possible. Alternatively, OX-40R might be up-regulated on transferred T cells at the tumor site, rendering them reactive to the mAb. Our study suggests OX-40R mAb to be a reagent of choice to augment T cell adoptive immunotherapy in clinical trials.     

Biological response modifiers (BRM) as antigens

In order to investigate tumoricidal effector cells in therapy by biological response modifiers (BRM) such asPropionibacterium acnes, bacillus Calmette-Guérin (BCG),Streptococcus pyogenes and a protein-bound polysaccharide (PSK), we established T cell lines specific for each BRM from BALB/c mice immunized with the corresponding BRM. These T cell lines proliferated and produced interleukin-2-(IL-2) and/or IL-4, but only in the presence of the relevant BRM and BALB/c spleen cells as the antigen and antigen-presenting cells respectively. Cross-functional experiments indicated that each BRM acts as a nominal antigen, but not as a non-specific immunostimulator. In addition, the T cell lines killed Ia-positive syngeneic B lymphoma cells, but only in the presence of the relevant BRM. These experiments excluded the possibility of cytotoxic effects by each BRM. The T cell lines and clones also killed Ia-negative bystander target cells, but only in the presence of both a relevant antigen and antigen-presenting cells. The T cell clones specific forS. pyogenes orP. acnes tested were Thy1⁺, L3T4⁺ and Lyt2^–. These results indicate that some BRM exert tumoricidal activity by inducing T cells that recognize them as an antigen and kill tumor cells in an antigen-specific manner. The T cells killed tumor targets in either a tumor-necrosis-factor(TNF)-dependent or a TNF-independent manner. The mediator of the latter pathway remains to be elucidated.     

Next-generation bromodomain inhibitors of the SWI/SNF complex enhance DNA damage and cell death in glioblastoma

Glioblastoma (GBM) is an aggressive brain cancer with a poor prognosis. While surgical resection is the primary treatment, adjuvant temozolomide (TMZ) chemotherapy and radiotherapy only provide slight improvement in disease course and outcome. Unfortunately, most treated patients experience recurrence of highly aggressive, therapy-resistant tumours and eventually succumb to the disease. To increase chemosensitivity and overcome therapy resistance, we have modified the chemical structure of the PFI-3 bromodomain inhibitor of the BRG1 and BRM catalytic subunits of the SWI/SNF chromatin remodelling complex. Our modifications resulted in compounds that sensitized GBM to the DNA alkylating agent TMZ and the radiomimetic bleomycin. We screened these chemical analogues using a cell death ELISA with GBM cell lines and a cellular thermal shift assay using epitope tagged BRG1 or BRM bromodomains expressed in GBM cells. An active analogue, IV-129, was then identified and further modified, resulting in new generation of bromodomain inhibitors with distinct properties. IV-255 and IV-275 had higher bioactivity than IV-129, with IV-255 selectively binding to the bromodomain of BRG1 and not BRM, while IV-275 bound well to both BRG1 and BRM bromodomains. In contrast, IV-191 did not bind to either bromodomain or alter GBM chemosensitivity. Importantly, both IV-255 and IV-275 markedly increased the extent of DNA damage induced by TMZ and bleomycin as determined by nuclear γH2AX staining. Our results demonstrate that these next-generation inhibitors selectively bind to the bromodomains of catalytic subunits of the SWI/SNF complex and sensitize GBM to the anticancer effects of TMZ and bleomycin. This approach holds promise for improving the treatment of GBM.     

Adoptive immunity in mice challenged with L1210/DTIC clones

Summary New antigenic specificities, not detectable on parental cells, have been induced by many investigators in mouse lymphomas by treatment with the antitumor agent 5(3,3-dimethyl-1-triazeno)imidazole-4-carboxamide (DTIC). The antigens are transmissible, after withdrawal of the drug treatment, as an inheritable character. The mechanism of induction, the molecular nature, and the number of the new antigenic specificities have not been completely elucidated. Four clones from murine leukemia L1210 isolated and expanded in vitro were treated in vivo with DTIC and the new sublines were studied in detail. The four drug-treated sublines studied exhibited strong immunogenicity since they were rejected by syngeneic animals. Immunosuppressed animals challenged with 10⁷ A/DTIC or P/DTIC cells were reciprocally protected by the adoptive transfer of spleen cells from donors that had rejected a lethal challenge of A/DTIC or P/DTIC clones. In a similar fashion, the adoptive transfer of spleen cells obtained from animals that had rejected the Q/DTIC or the R/DTIC clones protected immunosuppressed mice challenged with Q/DTIC or R/DTIC cells. No antitumor activity was observed in cross-protective schedules other than those indicated. It was been concluded that (a) the L1210 leukemia line does not have antigenic cells, (b) four DTIC-treated clone sublines were rejected by compatible hosts, and (c) two mutually exclusive sets of antigens were expressed in four antigenic clone sublines.Research supported in part by P.F.O. Contract Grant from C. N. R., Rome, Italy     

Cell-based immunotherapy of prion diseases by adoptive transfer of antigen-loaded dendritic cells or antigen-primed CD4+ T lymphocytes

Prion diseases are neurodegenerative conditions caused by the transconformation of a normal host glycoprotein, the cellular prion protein (PrPc) into a neurotoxic, self-aggregating conformer (PrPSc). TSEs are ineluctably fatal and no treatment is yet available. In principle, prion diseases could be attacked from different angles including: blocking conversion of PrPc into PrPSc, accelerating the clearance of amyloid deposits in peripheral tissues and brain, stopping prion progression in secondary lymphoid organs, reducing brain inflammation and promoting neuronal healing. There are many indications that adaptive and innate immunity might mediate those effects but, so far, the achievements of immunointervention have not matched all expectations. Difficulties arise from the impossibility to diagnose TSE before substantial brain damage, poor accessibility of the CNS to immunological agents, deep immune tolerance to self-PrP and short term effects of many immune interventions contrasting with the slow progression of TSEs. Here, we discuss two approaches, inspired from cancer immunotherapy, which might overcome some of those obstacles. One is vaccination with antigen-pulsed or antigen-transduced dendritic cells to bypass self-tolerance. The other one is the adoptive transfer of PrP-sensitized CD4⁺ T cells which can promote humoral, cellmediated or regulatory responses, coordinate adaptive and innate immunity and have long lasting effects.Key words: prion, TSE, dendritic cells, CD4⁺ T cells, cellular immunotherapy, vaccination, adoptive cell transfer     

The effect of biological response modifiers on chronic and latent murine cytomegalovirus infections

Host-mediated antiviral effect of 2 biological response modifiers (BRM), OK-432, and PS-K, against murine cytomegalovirus (MCMV) was evaluated in chronically or latently infected mice. In the early stage of chronic MCMV infection, the BRM-induced resistance was evidenced by decrease in infectious viruses replicated in the salivary glands and by augmented cytotoxic activity of the spleen cells against YAC-1 cells and MCMV-infected mouse embryonic fibroblasts (MEF). In the late stage of chronic MCMV infection, the BRM treatment did not eliminate MCMV from the mice, but did prevent exacerbation of MCMV infection in the salivary glands induced by administration of cyclophosphamide (CY). In mice latently infected by MCMV, BRM treatment suppressed CY-induced reactivation of MCMV in the salivary glands. It was suggested that the antiviral effect of BRM against MCMV in chronically or latently infected mice was based on activation of natural killer (NK) cells and cytotoxic T lymphocytes (CTL).     

Degron mediated BRM/SMARCA2 depletion uncovers novel combination partners for treatment of BRG1/SMARCA4-mutant cancers

Recent studies have highlighted that cancer cells with a loss of the SWI/SNF complex catalytic subunit BRG1 are dependent on the remaining ATPase, BRM, making it an attractive target for cancer therapy. However, an understanding of the extent of target inhibition required to arrest cell growth, necessary to develop an appropriate therapeutic strategy, remains unknown. Here, we utilize tunable depletion of endogenous BRM using the SMASh degron, and interestingly observe that BRG1-mutant lung cancer cells require near complete depletion of BRM to robustly inhibit growth both in vitro and in vivo. Therefore, to identify pathways that synergize with partial BRM depletion and afford a deeper response, we performed a genome-wide CRISPR screen and discovered a combinatorial effect between BRM depletion and the knockout of various genes of the oxidative phosphorylation pathway and the anti-apoptotic gene MCL1. Together these studies provide an important framework to elucidate the requirements of BRM inhibition in the BRG1-mutant state with implications on the feasibility of targeting BRM alone, as well as reveal novel insights into pathways that can be exploited in combination toward deeper anti-tumor responses.     

Effector mechanisms in cyclosporine A-induced syngeneic graft-versus-host disease. Role of CD4+ and CD8+ T lymphocyte subsets

Syngeneic graft-versus-host disease (SGVHD) is a T cell-mediated autoimmune disease occurring postsyngeneic bone marrow transplantation and the administration of the potent immunosuppressive agent, cyclosporine A. Paradoxically, cyclosporine A disrupts the immunologic homeostasis governing self-tolerance. Our studies using an adoptive transfer model attempted to identify effector mechanisms associated with the autoimmune disease. Both CD4+ and CD8+ splenic T cells isolated from autoimmune donors were required for the adoptive transfer of active disease into lethally irradiated secondary recipients reconstituted with normal bone marrow. Doses of more than 5 x 10(6) of nylon wool depleted splenocytes from autoimmune donors effectively transferred disease into lethally irradiated secondary recipients. Splenocytes that are T cell depleted or CD4(+)-enriched cells did not elicit disease upon adoptive transfer. Nylon wool fractionated CD8+ splenocytes also failed to adoptively transfer disease unless high doses (greater than or equal to 30 x 10(6)) were used. The disease transferred with the CD8+ subset presented as acute type SGVHD and was self-limiting. The recombination of the individually isolated T cell subsets not only restored but also enhanced immune reactivity upon adoptive transfer. Moreover, use of the recombined subsets resulted in progressive disease with the development of chronic type SGVHD. The titration of each subset to the other suggested that a minimal number of CD4+ T cells was required to potentiate the CD8+ autoreactive cells in vivo. Further analysis of the helper cell involved demonstrated that it had a CD4+ CD45r- phenotype, characteristic of an amplifying helper cell population. Administration of IL-2 did not substitute for CD4+ Th cells but yet amplified the activity of unfractionated cells or recombined subsets implicating the role of other factors in the pathogenesis of SGVHD. Delineation of the effector mechanisms involved in SGVHD is critical in determining the underlying events that trigger either the production of autoreactive cells or the perturbation of the regulation of these autoreactive cells, culminating in autoimmunity.     

MyoD can induce cell cycle arrest but not muscle differentiation in the presence of dominant negative SWI/SNF chromatin remodeling enzymes

Cell cycle arrest is critical for muscle differentiation, and the two processes are closely coordinated but temporally separable. SWI/SNF complexes are ATP-dependent chromatin-remodeling enzymes that have been shown to be required for muscle differentiation in cell culture and have also been reported to be required for Rb-mediated cell cycle arrest. We therefore looked more closely at how SWI/SNF enzymes affect the events that occur during MyoD-induced myogenesis, namely, cell cycle regulation and muscle-specific gene expression, in cells that inducibly express dominant negative versions of Brahma (BRM) and Brahma-related gene 1 (BRG1), the ATPase subunits of two distinct SWI/SNF complexes. Although dominant negative BRM and BRG1 inhibited expression of every muscle-specific regulator and structural gene assayed, there was no effect on MyoD-induced activation of cell cycle regulatory proteins, and thus, cells arrested normally. In particular, in the presence or absence of dominant negative BRM or BRG1, MyoD was able to activate expression of p21, cyclin D3, and Rb, all of which are critical for cell cycle withdrawal in the G1/G0 phase of the cell cycle. These findings suggest that at least one basis for the distinct mechanisms that regulate cessation of cell proliferation and muscle-specific gene expression during muscle differentiation is that SWI/SNF-mediated chromatin-remodeling enzymes are required only for the latter.     

Therapy of murine leukemia with cyclophosphamide and immune Lyt-2+ cells: cytolytic T cells can mediate eradication of disseminated leukemia

Several animal models have been developed in which the adoptive transfer of specifically immune syngeneic T cells has been shown to mediate the eradication of established tumors. In adoptive chemoimmunotherapy (ACIT) of disseminated FBL leukemia with cyclophosphamide and immune T cells, the major effector T cell has been shown to be a noncytolytic Lyt-1+2- T cell that mediates its therapeutic effect without the participation of CTL. Because studies in other models have suggested that CTL can mediate an anti-tumor effect, the efficacy of Lyt-2+ T cells rendered highly cytolytic before adoptive transfer in ACIT of disseminated FBL was examined. The results demonstrated that such CTL had a detectable but limited therapeutic effect in the treatment of FBL. Because this limited activity of transferred purified CTL might have reflected a requirement for helper T cells to produce IL 2 for promotion of the in vivo survival and proliferation of the CTL, the effect of administering IL 2 to tumor-bearing hosts after transfer of CTL was examined. A dose of IL 2 previously shown to induce in vivo proliferation of transferred T cells rendered CTL that were minimally effective alone curative in ACIT of FBL leukemia. Thus, either lymphokine-producing T cells or the lymphokines produced by these cells are necessary for the full expression of the in vivo therapeutic potential of CTL.     

The role of IL-12 in maintaining resistance to Leishmania major

IL-12p40 is required for the maintenance of resistance during Leishmania major infection. In this study, we addressed how IL-12 mediates this function. First, we demonstrated that both subunits of IL-12, p40 and p35, were required for continued resistance to L. major. Second, using IL-12, IL-4 doubly deficient mice, we investigated the possibility that IL-12 inhibits IL-4-induced outgrowth of Th2 cells that might compete with Th1 cells. We found that even in the absence of a Th2 response, IL-12 was still required to maintain resistance. Next, using adoptive transfer of Thy-1 disparate CD4(+) T cells from L. major-healed mice, we were able to show that the loss of a protective response in L. major-infected IL-12-deficient mice is linked with the loss of Th1 cells. In contrast, there was an equal recovery of CD4(+) Th1 cells from wild-type and IL-12-deficient mice when transferred into mice that were not challenged with L. major. The ability of Th1 cells to survive regardless of IL-12 levels in the absence of Ag stimulation was confirmed by adoptive transfer studies of CD4(+) Th1 cells from DO11.10 TCR transgenic mice. Taken together, these results indicate that, rather than modulating Th2 responses or optimizing IFN-gamma production, the critical role for IL-12 in maintaining cell-mediated immunity may be to prevent the loss of Th1 cells during a challenge infection.     

In vivo cyclophosphamide and IL-2 treatment impedes self-antigen-induced effector CD4 cell tolerization: implications for adoptive immunotherapy

The development of T cell tolerance directed toward tumor-associated Ags can limit the repertoire of functional tumor-reactive T cells, thus impairing the ability of vaccines to elicit effective antitumor immunity. Adoptive immunotherapy strategies using ex vivo expanded tumor-reactive effector T cells can bypass this problem; however, the susceptibility of effector T cells to undergoing tolerization suggests that tolerance might also negatively impact adoptive immunotherapy. Nonetheless, adoptive immunotherapy strategies can be effective, particularly those utilizing the drug cyclophosphamide (CY) and/or exogenous IL-2. In the current study, we used a TCR-transgenic mouse adoptive transfer system to assess whether CY plus IL-2 treatment rescues effector CD4 cell function in the face of tolerizing Ag (i.e., cognate parenchymal self-Ag). CY plus IL-2 treatment not only enhances proliferation and accumulation of effector CD4 cells, but also preserves the ability of these cells to express the effector cytokine IFN-gamma (and to a lesser extent TNF-alpha) in proportion to the level of parenchymal self-Ag expression. When administered individually, CY but not IL-2 can markedly impede tolerization, although their combination is the most effective. Although effector CD4 cells in CY plus IL-2-treated self-Ag-expressing mice eventually succumb to tolerization, this delay results in an increased level of in situ IFN-gamma expression in cognate Ag-expressing parenchymal tissues as well as death via a mechanism that requires direct parenchymal Ag presentation. These results suggest that one potential mechanism by which CY and IL-2 augment adoptive immunotherapy strategies to treat cancer is by impeding the tolerization of tumor-reactive effector T cells.     

Administration of agonistic anti-4-1BB monoclonal antibody leads to the amelioration of experimental autoimmune encephalomyelitis.

4-1BB, a member of the TNFR superfamily, is a costimulatory receptor primarily expressed on activated T cells. It has been shown that the administration of agonistic anti-4-1BB Abs enhances tumor immunity and allogenic immune responses. Paradoxically, we found that the administration of an agonistic anti-4-1BB mAb (2A) dramatically reduced the incidence and severity of experimental autoimmune encephalomyelitis (EAE). Adoptive transfer of T cells from such treated mice failed to induce EAE, whereas anti-4-1BB treatment following adoptive transfer of encephalitogenic T cells did not prevent EAE pathogenesis. These results suggest that anti-4-1BB treatment during the induction phase inhibits autoreactive T cell immune responses rather than preventing T cell trafficking into the CNS. This was substantiated by the observations that draining lymph node cells from anti-4-1BB-treated mice failed to respond to Ag stimulation in vitro. In addition, we found that such treatment initially promotes the activation and proliferation of Ag-specific CD4+ T cells but subsequently increases their probability of undergoing activation-induced cell death, thereby inhibiting effector T cell responses. More importantly, 2A treatment also inhibits the relapse of EAE in a clinically relevant murine model of multiple sclerosis. This study indicates that the agonistic Ab against 4-1BB can potentially be used as a novel immunotherapeutic agent for treating autoimmune diseases.     

Protective effect of biological response modifiers on murine cytomegalovirus infection

Pretreatment with two biological response modifiers (BRM), OK-432 and PS-K, protected mice from lethal infection by murine cytomegalovirus (MCMV). This was evidenced by an increase in 50% lethal doses and a decrease in titers of infectious viruses replicated in the liver and spleen. Spleen cells from the BRM-treated mice augmented the natural killer (NK) cell activity and suppressed the replication of MCMV in vitro. During MCMV infection, the NK cell activity of the spleen cells was maintained at a high level in the BRM-treated mice, whereas it was severely impaired in untreated mice. The BRM-induced protection was nullified by concomitant administration of antiasialo GM1 antibody. Interferon was neither induced by BRM treatment nor enhanced in BRM-pretreated and MCMV-infected mice. Thus, the protective effect of OK-432 and PS-K seems to be based on activation of NK cells and prevention of MCMV-induced inhibition of the NK cell activity.