Indolent aspergillus arthritis complicating fludarabine-based non-myeloablative stem cell transplantation.

Fungal arthritis and osteomyelitis are rare and documented mainly in immunocompromised or neutropenic patients. Patients receiving therapeutic immunosuppression for organ transplants have also reported to suffer from aspergillus osteoarthritis. We describe two patients with aspergillus arthritis of the knee joint following fludarabine-based non-myeloablative stem cell transplantation. Both were suffering from acute and chronic GVHD and treated with heavy immunosuppression including steroids and cyclosporine. Interestingly in one of our patients, the arthritis was almost asymptomatic and did not spread to other organs. Heavy pre- and post-transplant immunosuppression is a major risk factor for invasive fungal infection, which can involve remote organs and manifest in an indolent and atypical manner.

Amelioration of diabetes in nonobese diabetic mice with advanced disease by linomide-induced immunoregulation combined with Reg protein treatment.

Oral linomide, (quinoline-3-carboxamide), has been shown to prevent autoimmune insulitis, islet destruction, and diabetes in NOD mice treated at an early stage of the disease, but confers only partial protection in animals with advanced disease. Reg protein, the gene product of a complementary DNA isolated from a regenerating rat islet library, has been previously shown to induce expansion of beta-cell mass in pancreatectomized rats. To determine the effect of treatment combining immunomodulation and Reg protein on advanced autoimmune diabetes, we treated female NOD mice with oral linomide and i.p. Reg protein injections. In 14-week-old animals with less severe disease (glucose tolerant), treatment with each agent alone resulted in amelioration of diabetes, as did treatment with Reg alone in 5-week-old prediabetic mice. In 14-week-old animals with more severe disease (glucose intolerant), only treatment with the combination of both agents, but not that with each separately, resulted in amelioration of diabetes. Our study suggests that treatment aimed at abrogation of autoimmunity combined with expansion of beta-cell mass constitutes a potential therapeutic approach for treatment of insulin-dependent diabetes mellitus.

Nonmyeloablative stem cell transplantation and cell therapy as an alternative to conventional bone marrow transplantation with lethal cytoreduction for the treatment of malignant and nonmalignant hematologic diseases.

Myeloablative conditioning associated with hazardous immediate and late complications is considered as a mandatory first step in preparation for allogeneic blood or marrow transplantation (allogeneic BMT) for the treatment of malignant hematologic disorders and genetic diseases. Immune-mediated graft-versus-leukemia (GVL) effects constitute the major benefit of allogeneic BMT. Therefore, we have introduced the use of relatively nonmyeloablative conditioning before allogeneic BMT aiming for establishing host-versus-graft tolerance for engraftment of donor immunohematopoietic cells for induction of GVL effects to displace residual malignant or genetically abnormal host cells. Our preliminary data in 26 patients with standard indications for allogeneic BMT, including acute leukemia (n = 10); chronic leukemia (n = 8), non-Hodgkin's lymphoma (n = 2), myelodysplastic syndrome (n = 1), multiple myeloma (n = 1), and genetic diseases (n = 4) suggest that nonmyeloablative conditioning including fludarabine, anti-T-lymphocyte globulin, and low-dose busulfan (8 mg/kg) is extremely well tolerated, with no severe procedure-related toxicity. Granulocyte colony-stimulating factor mobilized blood stem cell transplantation with standard dose of cyclosporin A as the sole anti-graft-versus-host disease (GVHD) prophylaxis resulted in stable partial (n = 9) or complete (n = 17) chimerism. In 9 patients absolute neutrophil count (ANC) did not decrease to below 0.1 x 10(9)/L whereas 2 patients never experienced ANC < 0.5 x 10(9)/L. ANC > or = 0.5 x 10(9)/L was accomplished within 10 to 32 (median, 15) days. Platelet counts did not decrease to below 20 x 10(9)/L in 4 patients requiring no platelet support at all; overall platelet counts > 20 x 10(9)/L were achieved within 0 to 35 (median 12) days. Fourteen patients experienced no GVHD at all; severe GVHD (grades 3 and 4) was the single major complication and the cause of death in 4 patients, occurring after early discontinuation of cyclosporine A. Relapse was reversed by allogeneic cell therapy in 2/3 cases, currently with no residual host DNA (male) by cytogenetic analysis and polymerase chain reaction. To date, with an observation period extending over 1 year (median 8 months), 22 of 26 patients (85%) treated by allogeneic nonmyeloablative stem cell transplantation are alive, and 21 (81%) are disease-free. The actuarial probability of disease-free survival at 14 months is 77.5% (95% confidence interval, 53% to 90%). Successful eradication of malignant and genetically abnormal host hematopoietic cells by allogeneic nonmyeloablative stem cell transplantation represents a potential new approach for safer treatment of a large variety of clinical syndromes with an indication for allogeneic BMT. Transient mixed chimerism which may protect the host from severe acute GVHD may be successfully reversed postallogeneic BMT with graded increments of donor lymphocyte infusions, thus resulting in eradication of malignant or genetically abnormal progenitor cells of host origin.

Should soybean agglutinin purging be performed in breast cancer patients undergoing autologous stem cell transplantation? A retrospective analysis of 48 patients.

High-dose chemotherapy (HDC) followed by autologous stem cell transplantation (ASCT) has gained an increasing role in the treatment of high-risk Stage II-III and/or metastatic breast cancer patients. Several investigators reported on a high rate of tumor cells contaminating the bone marrow and peripheral blood stem cell collection. Nevertheless, the clinical implication of reinfusion of tumor cells with the stem cells to the relapse rate is still uncertain. In this retrospective analysis we compare the outcome and the toxicity of 29 patients with high-risk Stage II-III and 19 metastatic breast cancer patients who underwent HDC with ASCT. Thirteen patients underwent transplant with soybean agglutinin (SBA)-purged graft, while 35 consecutive patients received unmanipulated graft. Engraftment was significantly faster for the nonpurged transplant. No differences in disease-free survival, freedom from relapse, or overall survival were noted in both groups during a median follow up time of 14 months. We conclude that tumor cell purging using SBA in breast cancer patients is not warranted. New purging methods are needed to assess the role of tumor cell purging in breast cancer patients.

Immunomodulatory effects of linomide in animals immunized with immunopathogenic retinal antigens: dissociation between different immune functions.

Linomide (LS-2616, quinoline-3-carboxamide) has been reported to exert a diverse range of effects on the immune system. On one hand, this drug was found to stimulate the immune system and to enhance activities such as DTH or allograft rejection. On the other hand, linomide was shown to inhibit the induction of experimental autoimmune encephalomyelitis and myasthenia gravis, as well as the development of diabetes in non-obese diabetic (NOD) mice. Here we report the effects of linomide in animals immunized with uveitogenic retinal antigens. Treatment with linomide completely inhibited the development of experimental autoimmune uveoretinitis (EAU) in mice immunized with interphotoreceptor retinoid-binding protein and markedly suppressed EAU in rats immunized with S-antigen (S-Ag). In addition, linomide-treated rats exhibited reduced antibody production and lymphocyte proliferative response to S-Ag. In contrast to these suppressive activities, linomide treatment did not affect the development of adoptively transferred EAU in rats and moderately enhanced the DTH reactions to S-Ag in immunized rats in which EAU and other immune responses to this antigen were suppressed.

Modulation of experimental systemic lupus erythematosus with linomide.

OBJECTIVE: The objective of this study was to assess the beneficial effects of an early administration of low dose linomide, a new immunomodulator, in an animal model of experimental systemic lupus erythematosus (SLE). METHODS: Experimental SLE was induced in naive BALB/c mice, by immunization with anti-DNA mAb (MIV-7). Control Mice immunized with irrelevant human IgM served as controls. The immunized mice were treated with linomide (0.1 mg/ml in the drinking water), four weeks prior to the first immunization, at an early stage of the disease induction (one month after boost injection), or at a later stage (3 months following boost immunization). The treatment duration was 3 months in all schedules. The follow-up studies continued for 8 weeks after discontinuation of the treatment. The presence in the serum of autoantibodies against ssDNA, dsDNA histones, phospholipids and an irrelevant autoantigen-pyruvate dehydrogenase, was determined by enzyme-linked immunosorbent assay (ELISA). The clinical parameters assessed included erythrocyte sedimentation rate, peripheral blood cell counts and proteinuria. RESULTS: There was a 50-64% decrease in autoantibody levels in the sera of mice immunized with anti-DNA (MIV-7) mAb at the early stage of experimental SLE in mice which received linomide for a period of 3 months. No effect of linomide was noted in mice which received the drug during the later stages of experimental SLE when the disease was fully developed. Linomide had a preventive effect on the induction of experimental SLE in naive mice, when the treatment was initiated before the induction of the disease. This effect was abolished following cessation of the treatment. CONCLUSIONS: Linomide proved to be effective at the early stages of induction of the experimental SLE. However, the autoantibody levels rose following discontinuation of the therapy.

Differential killing of murine B-cell leukemia (BCL1) by photosensitization with merocyanine 540: implications for autologous bone marrow transplantation.

The efficacy of photosensitization by merocyanine 540 (MC540), a lipophilic fluorescent dye, was investigated in the murine B cell leukemia (BCL1). Normal BALB/c mice were injected with BCL1 cells exposed to MC540, followed by photosensitization with white light for 15 min to 2 h. Mice injected with BCL1 cells exposed for 1 or 2 h showed no sign of leukemia. Lethally irradiated mice were successfully reconstituted with mixtures of syngeneic bone marrow (BM) and BCL1 cells treated with MC540 following exposure to white light. Exposure of BM/BCL1 mixtures for 2 h proved to be effective in purging all BCL1 cells without affecting BM viability, as documented by normal hemopoietic reconstitution of all recipients surviving without evidence of leukemia. All recipients of untreated BM/BCL1 cell mixtures developed leukemia within 42 days. Adoptive transfer of 10(6) spleen cells obtained from treated mice into secondary naive syngeneic recipients was carried out in order to test for dormant BCL1 cells in treated recipients. No leukemia developed in any of the secondary recipients. Previous studies indicate that as few as 10, or possibly less, BCL1 cells are sufficient to cause lethal disease in BALB/c recipients. Our results suggest that MC540 may be an extremely potent tool for in vitro elimination of residual tumor cells while leaving uncommitted progenitor hemopoietic cells intact for hemopoietic reconstitution following lethal marrow ablation.