Immune "tolerance profiles" in donor bone marrow infused kidney transplant patients using multiple ex vivo functional assays.

Ex vivo identification of donor-specific unresponsiveness in organ transplant recipients is important for immunosuppression (IS) minimization. We tested three groups of stable living, related-donor kidney transplant patients up to 11 years postoperatively, i.e., 20 haploidenticals with donor bone marrow cell (DBMC) infusions, eight noninfused haploidentical controls (haplo controls), and 11 HLA-identical controls (HLA-id), using multiple ex vivo immune assays. We observed that no patients developed donor-specific antibodies. The majority showed donor-specific CTL unresponsiveness from year 1 onward. Thirteen of 20 DBMC recipients became specifically donor MLR nonreactive. Depletion of donor cells in DBMC recipients still MLR reactive increased donor-specific reactivity by 75% +/- 36% (p = 0.04). Adding them back in low concentration caused antigen specific inhibition. The frequencies of ELISPOT granzyme-B and interferon-gamma-producing cells somewhat paralleled the CTL and MLR responses. In the trans vivo DTH, 14 of 19 DBMC recipients demonstrated donor-specific unresponsiveness and 16 of 19 showed "linked suppression," vs none of eight and one of eight haplo controls and vs six of 10 and one of 10 HLA-ids, respectively. Most importantly, when all six assays were performed simultaneously, 10 of 18 DBMC, five of 10 HLA-ids, and no haplo controls were specifically donor unresponsive long term. We propose that a cluster analysis combining these assays will reveal tolerant recipients in whom IS minimization may safely be tested. This appears to have occurred in many DBMC-infused recipients.

Evaluation of the tolerogenic effects of donor bone marrow cells using a severe combined immunodeficient mouse-human islet transplant model.

The immunoregulatory role of human donor bone marrow cells (DBMC) has been studied extensively in our laboratory using in vitro and ex vivo assays. However, new experimental systems that can overcome the limitations of tissue culture assays but with more clinical relevance than purely animal experimentation, needed to be generated. Therefore we have developed a new human peripheral blood lymphocyte (PBL) severe combined immunodeficient (SCID) mouse islet transplantation model without the occurrence of graft-versus-host disease (GvHD) and have used it to evaluate the tolerogenic effects of DBMC. Nonobese diabetogenic (NOD)-SCID mice were transplanted with human deceased donor islets and were reconstituted with human PBL (allogeneic to islets; denoted as recipient) with or without DBMC from the islet donor. It was observed that the most cellularly economical dose was 3000 islets per animal and that injection into the portal vein was better than implantation under the kidney capsule. Even though maximal lymphoid reconstitution was observed with 40-million fresh and anti-CD3 activated recipient PBL (conventional method), the mice developed severe graft GvHD. However, with the new method of reconstitution where animals were injected with 20-million anti-CD3-activated plus 40-million anti-donor-activated recipient PBL, no discernible GvHD was observed. More importantly, this latter method was associated with islet transplant rejection, which in turn could be abrogated by co-injection of the animals with DBMC. These in vivo results confirmed our previous in vitro observations that human DBMC have regulatory activity.

Campath-1H induction therapy in African American and Hispanic first renal transplant recipients: 3-year actuarial follow-up.

BACKGROUND: In a retrospective study of the first 75 primary renal transplant patients given alemtuzumab induction at our center, 20 were African American (27%), 32 were Hispanic (43%), and 23 were non-African American, non-Hispanic (31%). METHODS: Alemtuzumab was given intraoperatively and 4 days later (0.3 mg/kg), with planned low-dose maintenance mycophenolate mofetil (500 mg twice daily) and tacrolimus (targeted trough levels of 5 to 7 ng/ml) and no corticosteroid therapy after the first week. Median follow-up among ongoing survivors with a functioning graft was 45 months. RESULTS: Three-year actuarial patient and graft survival rates were 95% and 85% in African Americans, 89% and 78% in Hispanics, and 96% and 96% in non-African Americans, non-Hispanics, respectively (not significant). Bioavailability of tacrolimus was significantly lower among African Americans in comparison with the other patient subgroups (P

Improved metabolic control and quality of life in seven patients with type 1 diabetes following islet after kidney transplantation.

BACKGROUND: The beneficial effects of glycemic control on both survival and function of transplanted kidneys in patients with type 1 diabetes mellitus (T1DM) and end-stage renal disease (ESRD) have been recognized. METHODS: Herein, we present the clinical outcome of a single-center pilot trial of islet after kidney (IAK) transplantation in seven patients with T1DM. The immunosuppression protocol for the kidney graft was converted to sirolimus+tacrolimus regimen 6 months before islet transplantation to exclude negative effects on kidney graft function. Primary endpoint was achievement of insulin independence after transplantation. Clinical outcome, metabolic control, severe hypoglycemia, kidney function, Quality of Life (QOL) psychometric measures, and adverse events were monitored. RESULTS: Seven patients showed graft function with improved metabolic control (A1c, fasting glycemia, and metabolic tests) after IAK (14,779+/-3,800 IEQ/kg). One-year insulin independence was 30% with persistent graft function in 86% (C-peptide-positive). A1c reduction was 1.95+/-0.31% from baseline (P<0.0001). No episodes of severe hypoglycemia were observed, even after resuming insulin. The direct consequence of these benefits was a significant improvement in diabetes QOL. Adverse events included procedure-related pleural effusion (n=2), cholecystitis (n=1), and additional immunosuppression-related, all resolved without sequelae. Kidney function (by estimated glomerular filtration rate) remained stable during follow-up in six of seven patients. CONCLUSIONS: Islet transplantation represents a feasible therapeutic option for patients with T1DM bearing a stable kidney allograft. Insulin independence at 1 year is lower than what reported in islet transplant alone. Nevertheless, clear benefits in terms of optimal metabolic control and absence of severe hypoglycemia are invariably present.

Allosensitization of islet allograft recipients.

BACKGROUND: The immune monitoring of islet transplant recipients includes the assessment of panel reactive antibodies (PRA). A negative association of PRA+ with allogeneic solid organ graft survival has been recognized, but scattered data is available for islet transplantation. METHODS: We performed a retrospective analysis of PRA status in 66 patients with type 1 diabetes mellitus recipient of islet allografts between 1985 and 2006. RESULTS: Pretransplant PRA+ was observed in 10 subjects in the old trials and associated with kidney transplantation and/or pregnancies. Thirteen subjects displayed PRA+ at follow-up, eight of whom were de novo. Overall, PRA+ did not correlate with islet graft outcome: long-term graft survival was observed in the presence of basal or persistent PRA+ and graft dysfunction occurred also in the absence of PRA+. Loss of graft function was associated with PRA+ after lowering of immunosuppression or after infection episodes. Loss of C-peptide did not affect kidney graft function even in simultaneous islet-kidney transplant recipients. Mostly, PRA remained negative under adequate immunosuppression. Patients whose immunosuppression was discontinued invariably developed PRA+. CONCLUSIONS: Monitoring of PRA under immunosuppression may have little clinical value under adequate immunosuppression in islet transplant recipients. The implications of allosensitization after discontinuation of immunosuppression need to be evaluated to define the real clinical impact in this patient population.

Blood citrulline level is an exclusionary marker for significant acute rejection after intestinal transplantation.

BACKGROUND: Serum citrulline is a marker for acute cellular rejection (ACR) after intestinal transplantation; however, its clinical utility has not yet been established. The goal of this study was to determine clearcut serum levels beyond which the diagnosis of acute rejection could be supported or refuted, and predictors of citrulline levels posttransplant from which more accurate estimates of sensitivity and specificity could be obtained. METHODS: Since March 2004, we obtained 2135 dried blood spot (DBS) citrulline samples from 57 intestinal transplant recipients at or beyond 3 months posttransplant. Stepwise linear regression was performed to determine the most significant multivariable predictors of the patient's DBS citrulline level. RESULTS: Seven characteristics were associated with a significantly lower citrulline in multivariable analysis: presence of mild, moderate, or severe ACR; presence of bacteremia or respiratory infection; pediatric age; and time from transplant to DBS sample (P<0.00001 in each case). Using a <13 vs. > or =13 micromoles/L cutoff point, the sensitivity for detecting moderate or severe ACR and the negative predictive value were high (96.4% and >99% respectively). Specificity was 54% to 74% in children and 83% to 88% in adults. CONCLUSIONS: Citrulline levels <13 micromoles/L should alert the clinical team that a serious problem (rejection or infection) could be looming in a previously stable intestinal recipient. Levels > =13 micromoles/L practically rule out moderate or severe rejection.

Inhibition of NF-kappa B during human dendritic cell differentiation generates anergy and regulatory T-cell activity for one but not two human leukocyte antigen DR mismatches.

We examined the in vitro inhibition of human monocyte-derived dendritic cells (DC) maturation via NF-kappaB blockade on T-cell allostimulation, cytokine production, and regulatory T-cell generation. DC were generated from CD14+ monocytes isolated from peripheral blood using GM-CSF and IL-4 for differentiation and TNF-alpha, IL-1beta, and PGE2 as maturational stimuli with or without the NF-kappaB inhibitors, BAY 11-7082 (BAY-DC) or Aspirin (ASA-DC). Stimulator and responder cells were one versus two HLA-DR mismatched in direct versus indirect presentation assays. Both BAY-DC and ASA-DC expressed high levels of HLA-DR and CD86 but always expressed less CD40 compared with controls. Some experiments showed slightly lower levels of CD80. Both BAY- and ASA-allogeneic DC and autologous alloantigen pulsed DC were weaker stimulators of T cells (by MLR) compared with controls, and there was reduced IL-2 and IFN-gamma production by T cells stimulated with BAY-DC or ASA-DC (by ELISPOT) (more marked results were always observed with ASA-treated DC). In addition, NF-kappaB blockade of DC maturation caused the generation of T cells with regulatory function (T regs) but only when T cells were stimulated by either allogeneic (direct presentation) or alloantigen pulsed autologous DC (indirect presentation) with one HLA-DR mismatch and not with two HLA-DR mismatches (either direct or indirect presentation). However, the T regs generated from these ASA-DC showed similar FoxP3 mRNA expression to those from nontreated DC. Extension of this study to human organ transplantation suggests potential therapies using one DR-matched NF-kappaB blocked DC to help generate clinical tolerance.

FoxP3 mRNA transcripts and regulatory cells in renal transplant recipients 10 years after donor marrow infusion.

BACKGROUND: We update more favorable 10-year deceased donor kidney transplant survival in 63 recipients infused perioperatively with donor vertebral body bone marrow (DBMC-i) vs. 219 noninfused controls having equivalent immunosuppression and demographics. We questioned if this was associated with putatively regulatory FoxP3 mRNA and cell phenotypes (CD4+CD25+high percentages and high DC2:DC1 ratios) in DBMC-i vs. noninfused controls. METHODS: Baseline studies were performed on peripheral blood lymphocytes (PBLs) vs. marrow in normal laboratory volunteers of CD4+CD25+high percentages and DC2:DC1 by flow cytometry, and FoxP3 mRNA in CD3+ cells by real-time polymerase chain reaction. Similar studies were performed on PBL of the majority of the 10-year patients remaining with graft function: 21 (of the remaining 37) DBMC-i vs. 55 (of the remaining 105) controls. RESULTS: In normal subjects, all parameters were significantly higher in marrow than in PBL, supporting our previous reports of ex vivo DBMC immunoregulation. At 9.8+/-.02 years posttransplant in DBMC-i vs. controls, death-censored percent graft failure was 17.5% vs. 32.9% (P=0.02) with 247.6+/-24 vs. 79.9+/-3.1 (mean+/-SE) FoxP3 copies/5,000 CD3+ cells (P=0.0001). PBL CD4+CD25+high percentages were lower, but DC2:DC1 values higher in both recipient groups than in end-stage renal disease patients who had lower FoxP3 levels (40.8+/-5.9, P<0.0001), consistent with non-CD4+CD25+high T regulatory cells generated long-term posttransplant. Individual higher FoxP3 values correlated with higher iliac crest chimerism in DBMC-i, but not in controls (with 50-fold lower chimerism). In chronically rejecting controls, FoxP3 was further decreased. CONCLUSIONS: Peritransplant DBMC-i has higher 10-year renal transplant acceptance, chimerism, and FoxP3 mRNA in thus-far unclarified regulatory cell phenotypes.

A novel approach to detect donor/recipent immune responses between HLA-identical pairs.

Renal transplant rejection and graft versus host reactions between HLA genetically-identical sibling (HLAgi) donor/recipient (D/R) pairs are thought to result from minor histocompatibility antigen (mHAg) disparities. We have compared two methods of measuring HLAgi D/R T lymphocyte responses to "matured" dendritic cells: 1.) a modified Cylex assay of CD4(+) ATP levels (MLDC-ATP) versus 2.) (3)H-thymidine uptake (MLDC-(3)H). The MLDC-ATP kinetics peaked at 48 hours versus the MLDC-(3)H at 7 days, and appeared more sensitive. We tested HLAgi (normal) volunteer siblings (NLs), and D/R sibling pairs before and after renal transplantation (pre-Tx and post-Tx). The overall frequencies of positive responses in the MLDC-ATP for HLAgi NLs, pre-Tx, and post-Tx D/R pairs were 63%, 50%, and 42%, respectively. The percentage with reciprocal responses was 37.5%, 20%, and 22.22%, respectively. In one set of three HLAgi (NLs) siblings (two males and one female), there was a nongender-associated differential response. There was no MLDC correlation with class I MHC-associated mHAg (SSP) incompatibility, nor could some MLDC positive reactive pairs theoretically process the necessary HLA-class I restriction molecules for presentation of known (nanomeric) mHAg peptides. Speculatively, the MLDC reflects class II MHC-restricted mHAg reactions (not yet definable), with possible effects of other polymorphic (nonhistocompatibility) immune response genes, and thereby may be a useful measurement of CD4(+) T-cell HLAgi transplantation immunity.

Induction of auto-reactive regulatory T cells by stimulation with immature autologous dendritic cells.

We have shown in ex vivo studies in donor bone marrow-infused kidney transplant recipients, that chimeric cells of either donor or recipient origin taken from the recipient's bone marrow down-regulated the recipient's cellular immune responses. In the present study, we have now induced regulatory T cells from peripheral blood mononuclear cells (PBMC) of renal transplant recipients or laboratory volunteers by multi-stimulation with autologous immature dendritic cell (iDC) enriched populations derived from either bone marrow cells (BMC) of the (immunosuppressed) kidney transplant recipients or PBMC of the laboratory volunteers (i.e., ibDC and ipDC, respectively). These regulatory T cells, induced by ibDC and ipDC, were autoreactive and designated as TAb and TAp with similar phenotypes and functional profiles. They were largely CD4 + CD25high, CD45RA low and CD45RO high, and uniformly expressed intracellular CTLA-4, and message of IL-4, IL-10, Foxp3, and differentially expressed TGFbeta. Their proliferative responses to autologous mature dendritic stimulating cells (mDC) were approximately two-fold stronger than to allogeneic mDC, and to allogeneic mDC were significantly lower than those of (control) autologous TPBL, suggesting an anergic state. TAb and TAp were not cytotoxic to autologous cells expressing Epstein-Barr virus (EBV) antigens, but were able to inhibit (regulate) the effector phase of this TPBL response to both autologous and allogeneic EBV lymphoblasts. This regulation appeared to require cell-to-cell contact.

Detection of HLA and MICA antibodies before kidney graft failure.

390 serum samples taken from a series of 28 patients who lost their grafts were retrospectively investigated for antibodies by single antigen HLA Class I&II and MICA Luminex beads. In 20 patients with immunological failure, 10 patients had DSA, 16 had epitope-related NDSA, and 16 had NDSA or MICA antibodies. In 16 which increased antibodies leading up to the graft failure, DSA were found in 7 patients. However, in 9 patients who similarly rejected their grafts, 6 were epitope-related DSA and 3 were NDSA. We were unable to establish why NDSA might be associated with the failure, but this was clearly the case in these patients. Of the 25 patients whose grafts were rejected, 23 (92%) produced antibody prior to failure. Our results strongly suggest that antibody monitoring is important as a means of detecting chronic rejection before a rise in SCr. Early detection of antibody would allow for appropriate and timely interventions to save a graft.

Utilization of dried blood spot citrulline level as a noninvasive method for monitoring graft function following intestinal transplantation.

BACKGROUND: Citrulline concentrations have been proposed as a marker for intestinal allograft rejection. We instituted dried blood spot (DBS) specimen monitoring of citrulline to simplify sample collection posttransplant. This study demonstrates the correlation between plasma and dried blood spot specimen citrulline concentrations after intestinal transplantation. METHODS: Plasma and DBS samples were analyzed by hydrophilic interaction chromatography tandem mass spectrometry. Comparison of the strength of linear correlation was made according to the type of surgery, sonication time, DBS citrulline levels, and the time interval between the blood sample collection and the assay date. RESULTS: A very strong linear correlation exists between the plasma and DBS citrulline concentrations (r=0.87; P<0.001). The correlation between plasma and DBS citrulline concentrations was maintained when evaluating only the intestinal transplant recipients. There was no significant difference in the strength of linear correlation according to sonication time, cirtrulline concentrations, or length of time to assay date. CONCLUSIONS: DBS citrulline monitoring will ease sample collection following intestinal transplantation and improve the ability to detect intestinal dysfunction and rejection by a noninvasive means.

A randomized trial of three renal transplant induction antibodies: early comparison of tacrolimus, mycophenolate mofetil, and steroid dosing, and newer immune-monitoring.

BACKGROUND: New trends in immunosuppression in clinical transplantation include the use of antibody induction agents in protocols that emphasize reduction or avoidance of steroids and calcineurin inhibitors. METHODS: In a randomized trial using three different antibody induction agents in 90 first renal transplant recipients from cadaver donors, group A received Thymoglobulin, group B received Alemtuzumab, and group C received Daclizumab. Maintenance immunosuppression included tacrolimus and mycophenolate in all three arms, and methylprednisolone in groups A and C only (standard clinical institutional practice). The targeted trough level of tacrolimus was between 8 and 10 ng/mL for groups A and C, respectively, with a targeted mycophenolate dose of 1 g twice daily. However, in group B, the target tacrolimus trough level was 4 to 7 ng/mL to reduce long-term nephrotoxicity, with 500 mg twice-daily doses of mycophenolate, without steroid maintenance. RESULTS: In this 15-month median postoperative interval report, there were no notable differences in demographics and patient and graft survivals. Acute rejection rates at 1 year were equivalent, that is, 5 of 30 in all three groups (16.6%). In group B, there was slightly lower renal function at 1 month, but no difference at 1 year. There was also significantly more leukopenia, but a greater percentage of T-regulatory cells and number of Fox-P3 mRNA copies by flow cytometry and semiquantitative polymerase chain reaction analysis, respectively, in group B. CONCLUSIONS: This preliminary analysis indicates that 80% of the patients in group B remained steroid-free 1 year postoperatively, with lower tacrolimus trough levels and no difference in other adverse events.

Serial ten-year follow-up of HLA and MICA antibody production prior to kidney graft failure.

The role of HLA antibodies in chronic allograft rejection was examined utilizing a unique resource of sera collected annually and stored over a 12-year period from patients with rejected or retained grafts. In patients selected for not having preformed HLA antibodies, 679 postoperative serial serum samples from 39 patients who rejected their grafts and 26 with functioning grafts were tested for HLA Class I and Class II antibodies by flow cytometry and for MICA antibodies by cytotoxicity on recombinant cell lines. HLA antibodies were found in 72% of patients who rejected grafts, compared to 46% with functioning transplants (p<0.05). In addition, the incidence of IgG HLA plus MICA antibodies was higher (77%) among those with failed transplants than those with functioning transplants (42%) (p<0.01). Finally, if patients with IgM anti-HLA antibodies were included, 95% of the 39 patients who rejected their grafts had HLA or MICA antibodies, compared to 58% with functioning grafts (p<0.01). Patients who rejected transplants had HLA and MICA antibodies more frequently than those with functioning grafts. These antibodies found in the peripheral circulation, were not necessarily donor-specific, but their association with failure is consistent with a causality hypothesis.

Islet transplantation in type 1 diabetes mellitus using cultured islets and steroid-free immunosuppression: Miami experience.

Following the success obtained with transplantation of fresh human islets under steroid-free immunosuppression, this trial evaluated the transplantation of islets that had undergone a period of in vitro culture and the potential of tumor necrosis factor (TNF-alpha) blockade to improve islet engraftment. Subjects included 16 patients with type 1 diabetes mellitus (T1DM); half were randomly assigned to receive Infliximab immediately preceding initial infusion. Immunosuppression consisted of daclizumab induction and sirolimus/tacrolimus maintenance. Out of 16 subjects 14 achieved insulin independence with one or two islet infusions; adverse events precluded completion in two. Without supplemental infusions, 11/14 (79%) subjects were insulin independent at 1 year, 6/14 (43%) at 18 months; these same subjects remain insulin independent at 33+/-6 months. While on immunosuppression, all patients maintained graft function. Out of 14 patients, 8 suffered chronic partial graft loss, likely immunological in nature, 5 of these received supplemental infusions. Currently, 11 subjects remain on immunosuppression, 8 (73%) are insulin independent, two with supplemental infusions. Insulin independent subjects demonstrated normalization of HbA1c, fructosamine and Mean Amplitude of Glycemic Excursions (MAGE) values. No clinical benefit of infliximab was identified. These results demonstrate that transplantation of cultured human islet allografts results in reproducible insulin independence in all subjects under this immunosuppressive regimen, comparable to that of freshly transplanted islets (Edmonton protocol).

Campath-1H does not alter bone marrow cell regulatory function.

We have previously reported in laboratory volunteers (in vitro) and renal transplant recipients (ex vivo) that bone marrow cells (BMC) are potent downregulators of the immune response. Also, the use of alemtuzumab (Campath-1H, C1H) for immunodepletion is associated with the most potent lasting effects yet seen on T-cell immunity in renal transplantation. We questioned whether the administration of C1H to kidney allograft recipients of donor bone marrow cell (DBMC) infusions would lead to stronger or weaker immunoregulatory effects. Human BMC depleted of T cells (nT-BMC) were either untreated or treated with C1H and rabbit complement and compared for their ability to downregulate autologous or allogeneic T-cell responses and to generate T regulatory (T reg) cells. The proliferative responses to anti-CD3 monoclonal antibody of T cells derived from cocultures with C1H-treated or untreated autologous nT-BMC were equally suppressed, i.e., an equivalent alteration in CD3 complex signaling, not regained by the addition of interleukin 2. Adenosine triphosphate levels were also markedly reduced in T cells both from C1H-treated and untreated nT-BMC cocultures. The ability of C1H-treated or untreated nT-BMC to suppress autologous T-cell cytotoxic function was also equivalent, with a marked, but equivalent, capacity to induce CD4/CD25(high) T regs from CD3(+) cells, which effectively downregulated cytotoxic T cells. To mimic the clinical infusion of DBMC into (allogeneic) recipients, peripheral blood mononuclear cells were also cultured with allogeneic C1H-treated and untreated nT-BMC. T cells derived from these cultures secondarily stimulated with the same-donor mature antigen-presenting cells exhibited suppressed cytotoxicity by 85% and 54%, respectively. These in vitro studies suggest that C1H does not abrogate BMC immunoregulation and thus may allow its lympho-depleting effect to be synergistic.

The role of donor bone marrow infusions in withdrawal of immunosuppression in adult liver allotransplantation.

We investigated the role of donor bone marrow cell (DBMC) infusions in immunosuppression withdrawal in adult liver transplantation. Patients enrolled were at least 3 years post-transplantation, with stable graft function. Forty-five (study group: G1) received DBMC, and 59 (control group: G2) did not. Immunosuppression was reduced by one third upon enrollment, by another third the second year of the study and was completely withdrawn the third year. Patient and graft survival were similar between the two groups. Although rejection episodes were significantly less in G1 the first 2 years of the study (35% vs. 57%, p = 0.016), there was no significant difference overall (74% vs. 81%, p = 0.14). Until February 2004, 20 patients, 10 in each group, were immunosuppression free for 1-3 years. Approximately 20% of long-term survivors of liver transplantation can successfully discontinue their immunosuppression. DBMC infusions, do not increase this likelihood.

TNF-alpha, TGF-beta, IL-10, IL-6, and INF-gamma alleles among African Americans and Cuban Americans. Report of the ASHI Minority Workshops: Part IV.

Point mutations or single nucleotide substitutions in the regulatory regions of cytokine genes may affect levels of cytokine expression and have been associated with acute and chronic rejection in organ transplantation, severity of graft-versus-host disease in hematopoietic stem cell transplants, and predisposition to autoimmune disorders. Because these cytokine variants have been studied primarily among Caucasians, we defined the alleles and frequencies of five cytokines among 691 unrelated, adult African Americans and 296 Cuban Americans in the American Society for Histocompatibility/National Institutes of Health Minority HLA Workshops. The genotypes of all cytokines, except for transforming growth factor (TGF)-beta among African Americans, were found to be in Hardy-Weinberg's equilibrium. Genotype frequencies among African American and Cuban American participants were compared with those of 75 North American Caucasian bone marrow donors and with published frequencies. Significant differences were observed in all comparisons except between Cuban and Caucasian Americans for alleles of interferon (IFN)-gamma, interleukin (IL)-6, and IL-10. The most notable differences were in genotype frequencies of African Americans compared with those of the two other populations. The frequency of the IFN-gamma genotype A/A, which is associated with low expression, was significantly higher in African Americans than in Caucasian or Cuban Americans (0.66 vs 0.37 and 0.26, respectively; p < 0.0001 for both comparisons). The high-expression G/G genotype for IL-6 was more than twice as prevalent among African Americans as among Caucasians and 1.5 times more frequent than among Cuban Americans (respective frequencies: 0.85 vs 0.38 and 0.49; p < 0.0001 for both comparisons). In African Americans, the frequency of the high-expression genotype for IL-10, GCC/GCC, was approximately half that of the frequency in Cuban and Caucasian Americans (0.10 vs 0.19 and 0.23, respectively; p < 0.0001, p = 0.004). Because levels of expression can affect inflammation and immune regulation, differences in cytokine allele frequencies between racial or ethnic groups may contribute to different incidences of autoimmunity and allograft rejection.

The use of Campath-1H as induction therapy in renal transplantation: preliminary results.

BACKGROUND: In an attempt to reduce both initial and long-term (nephrotoxic) calcineurin inhibitor maintenance dosage and totally eliminate maintenance corticosteroids, alemtuzumab (Campath-1H) was used as induction therapy in first cadaver and non-HLA-identical living donor renal transplantation. METHODS: Forty-four de novo renal allograft recipients were treated with Campath-1H (0.3 mg/kg) on days 0 and 4 postoperatively, preceded by methylprednisolone boluses. Maintenance target 12-hr tacrolimus trough levels of 5 to 7 ng/mL were operational from the outset as well as (reduced) mycophenolate mofetil dosage of 500 mg twice daily. No corticosteroids were planned to be given after the first week postoperatively. RESULTS: With a median follow-up of 9 (range, 1-19) months, patient and graft survival rates are each at 100%. Biopsy-proven acute rejection was diagnosed in four patients. Infections requiring hospitalization developed in four patients. Thirty-eight recipients remain without the need for long-term corticosteroid therapy. CONCLUSIONS: In an early assessment, the combination of Campath-1H, low dosing of tacrolimus and mycophenolate mofetil, and avoidance of maintenance corticosteroid use seems to be safe and effective for kidney transplant recipients. Long-term outcomes will be reported in the future.

Preliminary experience with alemtuzumab (Campath-1H) and low-dose tacrolimus immunosuppression in adult liver transplantation.

BACKGROUND: The administration of alemtuzumab (Campath-1H [C1H]; Berlex Laboratories, Montville, NJ) at transplantation prevents a vigorous immune response and is believed to allow a gradual engagement of the host immune system. We report our preliminary experience with C1H and tacrolimus (Tac) immunosuppression in adult liver transplantation. METHODS: We administered C1H and low-dose Tac to 40 adult recipients of cadaveric liver allografts between December 2001 and April 2003. A control group who met the same eligibility criteria consisted of 50 liver transplant recipients treated with our standard Tac and steroids protocol. RESULTS: Baseline characteristics and patient and graft survival were similar (P >0.15). The incidence of acute rejection was significantly lower during the first 2 months posttransplantation (P =0.002) and slightly lower overall in the study group versus the control group at 12 months (46% vs. 55%, P =0.12, log-rank test). Median time to rejection among those experiencing rejection was significantly longer in the study group versus control group (2.76 vs. 0.34 months, P =0.0007). The mean Tac dose, 12-hr trough level, and percentage of patients receiving maintenance steroids were significantly lower in the group receiving C1H and Tac (P <0.0001 during the first 3 months, P <0.05 thereafter), as were the mean creatinine levels (P <0.05) and incidence of nephrotoxicity (P =0.004, conversion from Tac to other agents). Finally, in the group receiving C1H/Tac, patients with an average Tac trough level less than 6.5 ng/mL during the first 2 months post-transplantation demonstrated a significantly higher rejection rate beyond that time (P =0.02). CONCLUSION: C1H and low-dose Tac seems to be at least as effective as our standard Tac and steroids regimen in preventing acute rejection in adult liver allotransplantation with less renal toxicity and less use of maintenance steroids.