Therapeutic Donor Lymphocyte Infusion Research Articles

Allogeneic B Cell Response to H-Y Minor Histocompatibility Antigens after Donor Lymphocyte Infusion Correlates with Disease Response.

Donor lymphocyte infusion (DLI) can induce remission in many patients who relapse after allogeneic hematopoietic stem cell transplantation (HSCT). We have previously demonstrated that male HSCT patients with female donors frequently develop high-titer antibody responses to H-Y antigens that correlate with disease remission. DLI administered 6 months after T cell depleted HSCT results in a five-fold increase in peripheral B cell numbers. We sought to determine whether allogeneic B cell responses develop after DLI. We expressed 5 recombinant H-Y proteins (DBY, UTY, ZFY, RPS4Y and EIF1AY) and developed sensitive ELISA to quantify the development of specific anti-HY antibodies. First, we studied prophylactic DLI. Twenty-six patients who received T cell depleted HSCT followed 5–7 months later by prophylactic CD8 depleted DLI were tested for H-Y antibodies pre-DLI and 6–12 months after DLI. No H-Y antibodies were detected in any of the pre-DLI serum samples. However, all 6 male HSCT patients with female donors (F→M HSCT) developed high-titer antibodies against at least one H-Y antigen after DLI. In contrast, only 1/20 of the other donor/recipient gender combinations (4 M→M, 8 F→F, 8 M→F) resulted in H-Y antibody (p<0.005). Thus, mHA disparity is required for the development of allogeneic B cell responses after DLI. This robust development of H-Y antibody in 6/6 F→M patients who received TCD transplantation and prophylactic DLI was significantly greater than 3/9 who developed H-Y antibodies after receiving the same TCD HSCT without DLI (p=0.03). This suggests that DLI augments allogeneic B cell responses after T cell depleted HSCT. To examine the effects of therapeutic DLI, we studied 24 F→M HSCT patients who relapsed 60 days to 15 years (median 704 days) after transplant and subsequently received either unmanipulated DLI (1−3x107 CD3+ cells/kg; n=12) or CD8 depleted DLI (3x107 CD4+ cells/kg; n=12). Only 2/24 had any H-Y antibody at the time of relapse. After DLI, 17/24 (71%) developed antibody to at least one H-Y antigen, and this correlated with complete remission after DLI (p<0.001). Disease progression continued in all 7 patients who did not develop H-Y antibodies, but 15 of 17 patients who developed H-Y antibodies also attained complete remission. H-Y antibodies developed rapidly and were detected as early as 26 days after DLI. Fifteen of 17 patients (88%) became H-Y antibody positive before 150 days after DLI. In our previous study assessing HSCT alone, only 3 of 38 (8%) developed H-Y antibodies before 150 days. Complete remission was attained with similar frequencies after both CD8 depleted DLI (7/12) and unmanipulated DLI (8/12). However, significant differences were noted in H-Y antibody responses by DLI type. In contrast to unmanipulated DLI, patients receiving CD8 depleted DLI developed high-titer antibodies (p=0.045) against multiple H-Y antigens (p=0.012). In summary, H-Y antibodies frequently develop in male patients after infusion of female donor lymphocytes and this allogeneic B cell response correlates with clinical response to DLI.H-Y Antibody Results in Male Patients with Female DonorsAny H-Y Antibody2 or more H-Y AntibodiesTCD HSCT + prophylactic CD8 depleted DLI6/6 (100%)4/6 (67%)TCD HSCT alone3/9 (33%)1/9 (11%)CD8 depleted DLI for relapse9/12 (75%)9/12 (75%)Unmanipulated DLI for relapse8/12 (67%)2/12 (17%)

Donor lymphocyte infusions in adult haploidentical transplant: a dose finding study.

Haploidentical transplantation has become a clinical option for patients lacking a compatible donor. However, patients are still referred at advanced stages and are usually heavily pretreated. This results in a high risk of toxicity, relapses and infections. We therefore started a donor lymphocyte infusion (DLI) dose-finding protocol, to try to improve both relapse rate and immunity reconstitution. In all, 12 consecutive patients were investigated. All had a refractory, some progressive, disease. Conditioning consisted of TBI, melphalan, ATG, fludarabine and CSA pretransplant. In four rapidly progressive patients, Ara-C had to be given 1 week preconditioning. The graft was T- and B-cell depleted with a fixed reinfused CD3 dose of 5 x 10(4)/kg. All patients engrafted before day 20. G-CSF was given from day 5 post-transplant and replaced with GM-CSF in the last three patients. Nonrelapse related mortality was 0/12 at 1 year. DLI were started at day 28 (3 x 10(4) CD3/kg) in the two first patients. This resulted in acute graft-versus-host disease (aGVHD) and chronic graft-versus-host disease (cGVHD) in both, but they did not relapse. The next dose was 1 x 10(4)/kg monthly for 3 months. This was well tolerated with only one grade I GVHD. Given the high relapse rate, we escalated doses (1, 3 and 10 x 10(4)/kg). This produced GVHD in all. We next moved, to GM-CSF and 1 x 10(4) CD3/kg monthly. Overall, 6/12 patients relapsed and received therapeutic DLI, starting at 1 x 10(5) CD3/kg with escalation every 2 weeks. We conclude that prophylactic DLI are feasible in adult haploidentical transplantation, without GVHD at a monthly dose of 1 x 10(4) CD3/kg. They result in faster CD4 recovery and a low rate of infections. The impact of GM-CSF remains to be further investigated. This scheme seems ideal for patients transplanted early in the course of their disease. In very bad prognosis patients, it remains insufficient to rapidly induce a GVL effect. Escalated doses are feasible but the price is aGVHD. Therapeutic DLI can be given at higher doses, depending on the time post-transplant. Haploidentical transplantation with low-dose DLI is a safe procedure that should be considered in all patients needing a transplant, but lacking a matched donor, early in the course of the disease.

Selective removal of alloreactive cells from haematopoietic stem cell grafts: graft engineering for GVHD prophylaxis.

One of the main goals in allogeneic bone marrow transplantation is the abrogation of graft-versus-host disease with the preservation of antileukaemia and antiviral activity. We have established a novel system for the selective removal of alloreactive lymphocytes from donor grafts while retaining an effective allogeneic response to third-party stimulator cells. Initial feasibility studies were done with unrelated HLA-mismatched pairs and then extended into the matched setting. Mononuclear cells from HLA-matched donors were cocultured with irradiated recipient cells prestimulated with cytokines (gamma-IFN and TNF-alpha) in a modified mixed lymphocyte culture (MLC). Alloreactive donor lymphocytes were identified by expression of CD69, an early activation marker and selectively removed by paramagnetic bead sorting. The remaining 'non-alloreactive' lymphocytes were tested in proliferative assays against the original matched recipient and to a third-party donor. A mean depletion of proliferative capacity to 11.5 +/- 9.9% of the original matched recipient response was achieved while the residual third-party response was largely preserved at 77.8 +/- 20.9% which should translate into improved immune reconstitution and preservation of antiviral activity. The non-alloreactive lymphocytes could also possess functional antileukaemia activity. Moreover, the alloreactive cells are easily recoverable in this selective T cell depletion strategy for cryopreservation and ready for immediate access as therapeutic donor lymphocyte infusions in cases of frank relapse post transplant.