• Ovid Technologies, Inc.

Sixty-five kidney transplantations performed across a non-current alloantibody-positive T cell crossmatch or an alloantibody-positive B cell crossmatch were studied retrospectively. The DTT crossmatch was used to discriminate between IgM and IgG donor-reactive antibodies. Subsequently the HLA specificity of donor-reactive IgG antibodies was determined in the MAILA assay. The first transplantations performed across a non-current positive T cell DTT crossmatch (IgG) were associated with poor graft survival, as only 5 of 11 (45%) transplants were functioning at 1 year. The HLA specificity of donor T cell reactive IgG antibodies appeared to determine the fate of the graft: only 2 of 7 (29%) patients with donor HLA class I-reactive antibodies had functioning grafts at 1 year, whereas all 3 patients with donor T cell-reactive antibodies, lacking HLA specificity, had functioning grafts. In 17 first transplantations, 15 grafts (88%) transplanted across an IgM-positive B cell crossmatch were functioning at 1 year. In 9 re-transplantations we found 6 grafts (67%) functioning at 1 year. B cell-reactive IgG antibodies, however, were associated with poor graft survival. In 7 first transplantations 2 grafts (29%) were functioning at 1 year, and in 17 re-transplantations 8 grafts (47%) were functioning at 1 year. For 19 patients the HLA specificity of donor B cell-reactive IgG antibodies was determined. Thirteen patients had HLA class II (-DR and/or -DQ)--specific antibodies; of these, 4 (31%) had a functioning graft at 1 year. Two of 3 (67%) patients with weak HLA class I--reactive antibodies and 2 of 3 (67%) patients with B cell--reactive IgG antibodies without HLA specificity had a functioning graft at 1 year. Although the number of cases analyzed is small, the following conclusions can be drawn: First, in general, the presence of donor HLA class I-, HLA-DR-, and HLA-DQ-reactive IgG antibodies is a contraindication to transplantation. However, under certain so-far-unknown conditions, transplantation across donor-reactive HLA specific IgG alloantibodies might be possible. Second, renal transplantation can be safely performed across B cell-reactive IgM antibodies. Third, donor-reactive IgG antibodies that do not recognize HLA do not seem to be harmful.

• Ovid Technologies, Inc.

Should HLA antigens that were mismatched in a renal transplant that failed be avoided in subsequent transplants? There were 890 retransplantations reported to the UCLA International Kidney Transplant Registry between 1985 and 1993 that had been performed in the face of a repeat HLA incompatibility. The 1- and 3-year regraft survival rates were 67% and 55%, respectively, for these retransplants, compared with 73% and 60% for 3220 regrafts with no HLA-A, -B, or -DR antigens mismatched twice (P = 0.030). When the repeat HLA-mismatched antigens were examined by locus, there was no difference in regraft survival comparing patients with no repeat HLA incompatibilities with those mismatched twice for HLA-A or -B antigens only, but there was a significant long-term decrease in survival of patients mismatched twice for HLA-DR antigens. The 377 patients mismatched twice only for HLA-A or -B antigens had 1- and 3-year regraft survival rates of 67% and 59%, respectively, compared with 65% (P = 0.289) and 50% (P = 0.025) for 281 patients with HLA-DR repeat mismatches only. Repeat mismatches for a combination of HLA-A or -B and -DR antigens resulted in 65% and 44% 1- and 3-year regraft survival in 129 patients. The half-lives for retransplants with repeat HLA class I, II, and I and II incompatibilities were 8, 6, and 4 years, respectively (P = 0.005). The data do not support preemptive avoidance of repeat HLA-A or -B incompatibilities. The crossmatch test excludes relevant mismatches. Repeated HLA-DR incompatibilities are not excluded by crossmatch tests and have a deleterious effect on long-term regraft survival. HLA-DR antigens mismatched in a previous failed transplant should be avoided.

• Ovid Technologies, Inc.

Renal transplantation in patients presenting end-stage renal failure can be hampered by the presence of alloantibodies against HLA antigens. In 4 out of 5 patients with HLA-specific alloantibodies waiting for a renal allograft, treatment with high-dose i.v. Ig resulted in a prolonged suppression (over 3 months) of most of the panel-reactive anti-HLA antibodies (PRA). Intravenous polyclonal human Ig (IVIg) and F(ab')2 fragments from IVIg inhibited the binding of patients' plasma and IgG fractions to peripheral blood lymphocytes from normal donors as well as their cytotoxicity, suggesting that the in vivo effect of IVIg was mediated by the presence, in the IVIg preparation, of anti-idiotypes directed against idiotypes borne on the anti-HLA antibodies. Thus, treatment with IVIg can be a valuable tool toward the transplantation of immunized patients.

• Ovid Technologies, Inc.

This study was conducted to determine the efficacy of the T cell flow cytometry crossmatch (T-FCXM) test in 841 first cadaver donor transplants. Results showed one-year graft survival rates were 82% for T-FCXM-negative patients, compared with 75% for T-FCXM-positive patients (P = 0.01). Early one-month graft failure was 13 percentage points higher in those with a positive T-FCXM than those with a negative T-FCXM. The positive crossmatch patients also had more frequent immunological failures. A positive T-FCXM was found in 39% of the sensitized patients (PRA > 10%) and 8% of those who had not been sensitized. Patients with a positive T-FCXM in either category had a 74% graft survival rate. Thus, most of the T-FCXM-positive results occurred in patients with complement-fixing antibodies. It is suggested that flow cytometry crossmatching (FCXM) be used prospectively, despite the fact that many patients with a positive crossmatch did have successful transplants (TXs). In the current climate of a cadaver kidney scarcity and large recipient waiting pools, utilization of kidneys for patients with the highest probability of success seems a most prudent policy.

• Ovid Technologies, Inc.

A high level of panel-reactive antibodies (PRA) in potential renal transplant recipients is associated with a long waiting time until transplantation and correlates inversely with graft outcome. We report our experience with the employment of immunoadsorption (IA) using a column composed to sepharose-bound staphylococcal protein A (which has a relatively selective affinity for binding IgG compared with other immunoglobulins) to decrease the PRA levels and expedite transplantation in 6 highly sensitized potential renal transplant recipients (1 primary and 5 awaiting second transplants). All patients had PRA levels of greater than or equal to 70% for a duration of 1 year prior to IA. Only patients with antibody specificity localized to 1 or 2 HLA A or B antigens were accepted for the study. IA procedures were performed on alternate days until a twofold decrease in antibody titer had occurred (maximum: 6 procedures). Repeat procedures were initiated if the HLA antibody titer returned to its baseline value. Intravenous cyclophosphamide (CY) (10 mg/kg/day every 3 weeks) and methylprednisolone (MP) (0.5 mg/kg/day) were provided as adjunctive immunosuppression until transplantation. A total of 44 immunoadsorption procedures were performed (27 primary and 17 repeat) with treatment of 2.49 +/- 0.02 plasma volumes per session. Serum IgG concentration decreased 95 +/- 3% and PRA activity decreased 75 +/- 16% after the primary treatment course. Four patients received cadaveric grafts within 3.7 +/- 1.2 months following the last IA procedure. Three grafts are functioning at 1 year, 8 months, and 8 weeks posttransplant. The remaining graft demonstrated primary nonfunction. All four patients had a past positive crossmatch using pre-IA sera with their respective donors. Patients not transplanted exhibited rapid resynthesis of IgG and a return of the PRA towards baseline levels within a few weeks after IA. We conclude that IA can effectively remove HLA antibodies and expedite graft availability in highly sensitized patients.

• Ovid Technologies, Inc.

In an attempt to reduce anti-HLA immunization in 15 patients awaiting for renal grafts and who were immunized against 65% of a panel of lymphocytes (titre 1/8 to 1/128), were given 5 to 7 sessions of immunoadsorption on protein A columns, immunosuppressor drugs (corticosteroids: 1 mg/kg/day + cyclophosphamid: 2 mg/kg/day) and intravenous polyclonal immunoglobulins. The antibody titres decreased in all patients, but this protocol did not sufficiently block resynthesis of antibodies. Among the 12 patients who were transplanted, the graft functioned correctly in 8 after a follow-up of 3 months to 3 years. Three early graft failures occurred in the group of 5 patients whose had had a positive cross-match before treatment. This treatment did not appear to increase the frequency of infectious complications immediately after transplantation.

Antibodies against human lymphocyte antigens (HLA) are frequently seen among patients undergoing repeated renal transplantations. Graft survival can be improved by eliminating these antibodies by plasmapheresis before transplantation. In this study, we have tried a new extracorporeal procedure to remove the anti-HLA antibodies. An immunoadsorption column (IM-TR) with a matrix of polyvinyl alcohol (PVA) gel conjugated with a hydrophobic amino acid tryptophan was utilized. Previous results have shown that repeated IM-TR treatments are at least equally effective as plasmapheresis in reducing levels of specific immunoglobulins in treated patients. In this study, 7 HLA-immunized patients were treated before renal transplantation. Each patient was subjected to a total of 12 treatment sessions divided into 3 sessions per week. After each treatment session, the reduction of the immunoglobulins was less than what has been reported for plasmapheresis. This suggests that mechanisms other than immunoglobulin depletion are involved in the reduction of the total immunoglobulin levels. The IM-TR treatment resulted in a strong complement activation triggered by the alternative pathway. Since the adsorbed plasma was returned to the patient, exceedingly high levels of the activation fragment C3d (C3dg) were found in plasma during and after the treatment. We conclude that the extensive generation of C3dg may be one of the factors that plays a role in the reduction of the antibody levels since the C3dg fragment has been shown to down-regulate the immune response.

• Wiley