Eleanor L. Ramos

The evaluation of candidates for renal transplantation. The current practice of U.S. transplant centers.

The criteria for acceptance of candidates for renal transplantation varies throughout the United States. The Patient Care and Education Committee of the American Society of Transplant Physicians conducted a survey of all U.S. centers that participate in the United Network for Organ Sharing (UNOS) concerning their evaluation of adult candidates for kidney transplantation. The response to each question was examined according to the specialty of the individual who filled out the questionnaire, as well as the type of transplant center (university or private) and the size of the center. The response rate to the survey was 81% (147/182). We found the following: (1) university-based and larger centers accepted more medically complicated patients; (2) 83% noted that attendance to dialysis was an important indicator of compliance after transplantation; (3) 79% did not require preoperative blood transfusions for cadaver kidney recipients; (4) 66% set no specific upper age limit for transplantation; (5) 56% excluded patients with chronic active hepatitis in the setting of hepatitis B antigenemia; (6) 50% had no specific policy for evaluating hepatitis C antibody-positive patients, while 54% excluded the use of hepatitis C antibody-positive donors, and (7) 15% obtained coronary angiography on all diabetic patients. U.S. transplant centers have a heterogeneous approach to the evaluation of patients for renal transplantation, particularly in the areas of viral hepatitis, cardiovascular disease, and noncompliance. University-based centers and centers that perform a larger number of transplants accept more medically complicated patients.

Results of 3-year phase III clinical trials with daclizumab prophylaxis for prevention of acute rejection after renal transplantation

BACKGROUND Daclizumab (Zenapax, Roche Pharmaceuticals), a humanized monoclonal antibody directed against the alpha chain of the interleukin 2 receptor, has been shown to reduce the incidence of acute rejection at 6 months after renal transplantation in two phase III clinical trials. This report presents the combined 1- and 3-year outcomes of kidney transplant recipients who participated in these two phase III clinical trials. METHODS Data from two multicenter, randomized, placebo-controlled trials were evaluated with regard to graft survival, patient survival, incidence of malignancies (including lymphoma), renal function (serum creatinine and glomerular filtration rate [GFR]), and current maintenance immunosuppressive regimen. In addition, the impact of acute rejection and acute rejection requiring treatment with antilymphocyte therapy upon 3-year graft survival was evaluated. Daclizumab was compared to placebo on a background of cyclosporine (CsA), azathioprine, and corticosteroids (triple therapy, TT) or CsA and corticosteroids (double therapy, DT). RESULTS Treatment with daclizumab in the pooled analysis demonstrated a significant reduction in the incidence of biopsy-proven acute rejection episodes at 12 months posttransplant (43% vs. 28%, P<0.001). The 3-year graft survival was not significantly different between placebo and daclizumab-treated patients in the TT trial (83% vs. 84%) or in the DT trial (78% vs. 82%). Pooled patient survival was excellent in both placebo- (91%) and daclizumab- (93%) treated patients. The incidence of malignancies or posttransplant lymphoproliferative disorder (PTLD) in placebo- versus daclizumab-treated groups was comparable in both clinical trials. Renal function was similar between placebo- and daclizumab-treated groups in both the TT and DT trials. The occurrence of delayed graft function, acute rejection requiring antilymphocyte therapy at 6 months, and acute rejection at 12 months posttransplant were associated with decreased graft survival rates at 3 years posttransplant. CONCLUSIONS The beneficial effect of daclizumab prophylaxis upon the incidence of acute rejection after renal transplant with TT or with DT was not associated with adverse clinical sequelae, including the development of PTLD, at 3 years posttransplant. There was no beneficial effect of daclizumab on graft survival at 3 years, but the trial was inadequately powered to detect this. Both studies showed excellent graft and patient survival at 3 years.

A preliminary report of diltiazem and ketoconazole. Their cyclosporine-sparing effect and impact on transplant outcome.

A prospective randomized trial was conducted to compare the effect of diltiazem (DILT) with ketocon-azole (KETO) on sparing of cyclosporine dose and renal transplant outcome. Renal allograft recipients 18 years old and older were eligible for the study. Triple immunosuppression (TRIPLE) including prednisone, azathioprine, and CsA was administered to all patients. The maintenance CsA dose varied by study group. Patients were randomized to receive one of three treatment strategies: group 1—TRIPLE (CsA 8 mg/kg/day); group 2—TRIPLE (CsA 6 mg/kg/day) + DILT (60 mg b.i.d.); group 3—TRIPLE (CsA 3 mg/kg/ day) + KETO (200 mg/day). Modification of the DILT dose was allowed as needed to effect blood pressure control in group 2 patients. Mean 1-month CsA dose reductions were 30% and 60% of controls in group 2 and 3, respectively. A continued effect over time was observed in patients administered KETO but not DILT. At 1 year patients taking KETO required an average of 77% less CsA than the average dose necessary to effect similar parent CsA blood levels when no enzyme inhibitor was used. The use of KETO and DILT for 1 year allowed for 53% and 14% reductions in CsA cost, respectively. These savings include the cost of the KETO or DILT. Serum creatinines, mean arterial pressure (MAP), and incidence of liver function abnormalities were similar throughout treatment groups. The rate of rejection, time to rejection onset, and survival (GS/PS) were not different among the groups. Fungal infections were fewer in patients treated with KETO (12%) than in controls (16%) and patients randomized to DILT (19%). KETO failed to prevent Aspergillus infection in one individual. The investigation failed to identify any harmful result of treating renal allograft recipients with either DILT or KETO for the purpose of reducing CsA expense.

Thromboxane synthase expression in renal transplant patients with rejection

Thromboxane synthase (TS) catalyzes the formation of thromboxane (TxA2) in monocytes/macrophages, platelets, and various tissues. TxA2 is likely to play a role in graft dysfunction due to its vasoconstrictive and platelet aggregatory properties. We studied the expression of TS in 7 normal native kidneys, 29 consecutive renal allograft biopsies (performed for rising serum creatinine, n = 23, and delayed graft function, n = 6), and one transplant nephrectomy specimen with severe acute rejection. TS expression was determined by immunocytochemistry using a monoclonal antibody against human TS, Kon-7. Histologic grading of the transplant biopsy specimens was based on the Banff classification. The degree of TS staining was graded in the glomeruli, interstitium, tubules and vessels from 0 to 3+. Of 29 biopsies, 13 had chronic nephropathy (CN), 6 had acute rejection (AR) with chronic nephropathy (AR/CN), 4 had acute rejection (AR), and 6 had acute tubular necrosis (ATN). TS staining of native kidneys showed sporadic interstitial cells. The biopsy and transplant nephrectomy specimens showed significant staining, predominantly in the glomeruli and interstitium. Positively staining cells appeared to be of macrophage/monocyte lineage by morphology. The mean glomerular staining grade was significantly increased in specimens with AR (2.3 +/- 0.9) and the mean interstitial staining was increased in specimens with AR/CN (2.2 +/- 0.9). Follow-up renal function 6 months post-biopsy showed that patients with higher TS staining grades had a faster decline in graft function. In conclusion, TS expression is increased in patients with acute rejection with or without chronic nephropathy and is associated with more rapid deterioration in function.

Daclizumab (humanized anti-IL2Ralpha mAb) prophylaxis for prevention of acute rejection in renal transplant recipients with delayed graft function.

Background. The purpose of this retrospective study was to determine the benefits of daclizumab, (Zenapax®, Roche Pharmaceuticals) a humanized anti-interleukin-2R&agr; (IL-2R&agr;) monoclonal antibody, for prevention of acute rejection in renal transplant recipients with delayed graft function (DGF). Methods. Data from two multicenter randomized placebo-controlled trials were pooled. DGF was defined by urine output <30 cc/hour, decline in serum creatinine of <0.5 mg/dl, or the need for dialysis within the first 24 hours after transplantation. Results. At one year posttransplantation, the incidence of biopsy-proven acute rejection in patients with DGF was reduced from 44% in the placebo group to 28% in the daclizumab group. (P =0.03) Prophylaxis with daclizumab also delayed the onset of the first biopsy-proven acute rejection episode in patients with DGF from 29±43 days in the placebo group to 73±70 days in the daclizumab group. (P =0.004) The graft survival rates in patients with DGF at 1 year posttransplantation were 78% in the placebo group and 82% in the daclizumab treated group. (P =ns) Three patients in the placebo-treated group with DGF experienced graft loss due to acute rejection, whereas no patients in the daclizumab-treated group with DGF had graft loss due to acute rejection. The 1-year patient survival rate in those with DGF in the placebo and daclizumab groups were 93% and 98%, respectively. (P =ns) Conclusions. Daclizumab effectively reduced the incidence and delayed the onset of biopsy-proven acute rejection in this high-risk subgroup of patients with DGF after renal transplantation. Graft and patient survival rates were similar between placebo- and daclizumab-treated patients with DGF.

Differential IL-2 receptor expression in renal allograft recipients treated with an anti-IL-2-receptor antibody.

Patients were entered into a randomized trial of prophylaxis for renal allograft rejection by the administration of an anti-human IL-2 receptor antibody, anti-Tac, during the first ten days posttransplant. Interleukin-2 receptor (IL-2 R) expression was measured using two anti-IL-2 R monoclonal antibodies (moAbs), anti-Tac and 1HT4-4H3. These two antibodies recognize closely spaced epitopes on the 55 kD chain of the IL-2 R. IL-2 R expression was examined on peripheral blood small lymphocytes in three groups of patients who received: (A) cyclosporine CsA and prednisone for baseline immunosuppression (n = 9); (B) anti-Tac with CsA and prednisone as baseline immunosuppression (n = 12); and (C) anti-Tac with azathioprine and prednisone as baseline immunosuppression (n = 5). We found that large numbers of T cells express IL-2 receptors despite the presence of anti-Tac (average of IL-2 R-positive cells at day of peak IL-2 R expression 56.0 +/- 20.8% in group A, 65.2 +/- 26.6% in group B, 21.0 +/- 7.4% in group C). IL-2 R expression did not correlate with clinical activity, and the presence or accessibility of epitopes on the same 55 kD chain varied dramatically from patient to patient.

Ten-year experience with cyclosporine as primary immunosuppression in recipients of renal allografts.

Cyclosporine has been used as primary immunosuppression in renal allograft recipients in our unit for the past decade. The overall clinical experience and long-term effects of the agent are reviewed. There were 461 consecutive recipients of kidney grafts; 379 received grafts from cadaver donors (CAD) and 82 from living related donors (LRD). Four separate clinical protocols were used sequentially using progressively decreasing doses of CyA; azathioprine was added in group 4 recipients of LRD grafts, and in patients receiving secondary CAD grafts (group 5). The patient mortality rate was less than 5%, with sepsis being the prime contributor. The majority of kidney grafts were lost within the first 2 months after operation; those that never functioned were found almost invariably to have been irreversibly rejected. During the subsequent years of follow-up, attrition of CAD grafts was significantly greater than LRD grafts. In contrast, the attrition rate of primary and secondary CAD grafts was the same after the first 3 months, emphasizing the importance of early immunologic graft destruction. Primary nonfunction occurred in 49% of CAD kidneys and 17% of LRD grafts; however 71% of initially nonfunctioning LRD grafts never functioned at all compared to 34% of CAD grafts, the majority of such organs undergoing fulminate rejection. Individual graft loss after 1 year was almost inevitably due to chronic rejection; there were no differences in long-term allograft function among the treatment groups. Although CyA has improved overall results of kidney transplantation, chronic rejection remains a major unresolved problem.

Decrease in phenotypically defined T helper inducer cells (T4+4B4+) and increase in T suppressor effector cells (T8+2H4+) in stable renal allograft recipients

Two monoclonal antibodies, anti-2H4 and anti-4B4, reciprocally divide the T4+ (CD4+) and T8+ (CD8+) lymphocytes into T4+2H4+, T4+4B4+, T8+2H4+ and T8+4B4+ subsets. The T4+2H4+, T4+4B4+ and T8+2H4+ subsets possess suppressor-inducer, helper-inducer, and suppressor-effector function, respectively, as previously defined in a system of B cell immunoglobulin production. Using monoclonal antibodies, including anti-2H4 and anti-4B4, and flow cytometry, we monitored lymphocyte subpopulations in 66 renal allograft recipients. We found that patients with stable allograft function have a decrease in the percentage of total T4+ lymphocytes from 41.9 +/- 9.5% pretransplant (pre-Tx) to 36.3 +/- 13.9% posttransplant (post-Tx) (P less than 0.05). This decrease was seen mainly in the T4+4B4+ or helper-inducer subset from 20.8 +/- 4.7% (pre-Tx) to 16.0 +/- 6.3% (post-Tx) (P less than 0.005). Patients with stable function were also noted to have an increase in the percentage of total T8+ lymphocytes from 21.3 +/- 10.7% (pre-Tx) to 30.9 +/- 15.4% (post-Tx) (P less than 0.02). Examination of T8 subsets revealed that a statistically significant increase was seen in the T8+2H4+ or suppressor effector subset from 15.5 +/- 9.2% (pre-Tx) to 21.5 +/- 10.2% (post-Tx) (P less than 0.01). Additionally, serial studies on 14 patients revealed an increase in the %T4+2H4+ suppressor-inducer subset from 9.31 +/- 3.64% (pre-Tx) to 15.71 +/- 6.41% (post-Tx) (P less than 0.0025). Since the role of these subsets has not been established in alloimmunity, in vitro allogeneic studies of 2H4-enriched (2H4+) and 2H4-depleted (2H4-) lymphocytes from normal peripheral blood were performed. In the mixed lymphocyte reaction, 2H4+ cells proliferated less than 2H4- cells (cpm ratio 2H4+/2H4-: 0.63-0.84), but 2H4+ cells generated twice as much suppressor activity as 2H4- cells (ratio % suppression 2H4+/2H4-: 1.9-2.3). These results suggest that 2H4+ cells play a role in the suppressor limb of the alloimmune response and that the increase in cells of this phenotype in our transplant population may be responsible for the maintenance of stable allograft function.

Antigen specificity of mixed lymphocyte response-induced suppressor cells

The nature of the antigens recognized by mixed lymphocyte response-generated suppressor cells is currently unknown. Previous investigations have yielded conflicting results, with different studies finding that suppressor cells recognize HLA class I antigens, class II antigens, or neither. To characterize the antigens recognized by suppressor cells (modulators) further, we generated 36 different modulators and assayed them for suppressor activity against a random 48-member HLA-typed panel in a total of 473 assays. Logistic regression analysis of the data revealed that suppression was correlated with B and DQ antigenic sharing between the original stimulator (used to generate the suppressor cells) and the test culture stimulator (p = 0.0043 and 0.0277, respectively). A role for DR antigen sharing could not be excluded. Overall, 35% of all suppressed assays could not be accounted for by the sharing of either any classical private HLA antigens, or of HLA-A or B locus cross-reactive group specificities. Suppression in these instances may involve the sharing of minor antigenic determinants, unidentified private HLA epitopes, or possibly another gene related to suppression that exists in linkage disequilibrium with the HLA-B locus or the DQ subregion.

T Cells Marked by the 2H4 Antigen Function in Allosuppression

A mouse monoclonal antibody (MAb), anti-2H4 (IgG1 subclass), has recently been described (1) that recognizes 200/220 kD glycoproteins (2) of the leukocyte common antigen/T200 family. The 2H4 antigen is found on 42% of unfractionated human T cells, 41% of CD4+ lymphocytes, 54% of CD8+ lymphocytes, and over 30% of both peripheral blood B cells and null cells. In a pokeweed mitogen system that measures B cell immunoglobulin production, the CD4+2H4+ cells were found to be inducers of suppression (1) and CD4+2H4− cells to be inducers of help (3). In the autologous mixed lymphocyte response (AMLR), CD8+2H4+ cells were found to have suppressor effector function (4). Additionally, in a concanavalin A− activated system, suppressor cell activity belonged to the 2H4+ subset of T cells (2). As the role of these subsets have not been established in alloimmunity, we studied the proliferative response and generation of suppressor cells in an allogeneic MLR using 2H4 enriched or depleted cells as the responding population.