Selective Bcl-2 inhibition promotes hematopoietic chimerism and allograft tolerance without myelosuppression in nonhuman primates.

Hematopoietic stem cell transplantation (HSCT) has many potential applications beyond current standard indications, including treatment of autoimmune disease, gene therapy, and transplant tolerance induction. However, severe myelosuppression and other toxicities after myeloablative conditioning regimens have hampered wider clinical use. To achieve donor hematopoietic stem cell (HSC) engraftment, it appears essential to establish niches for the donor HSCs by depleting the host HSCs. To date, this has been achievable only by nonselective treatments such as irradiation or chemotherapeutic drugs. An approach that is capable of more selectively depleting host HSCs is needed to widen the clinical application of HSCT. Here, we show in a clinically relevant nonhuman primate model that selective inhibition of B cell lymphoma 2 (Bcl-2) promoted hematopoietic chimerism and renal allograft tolerance after partial deletion of HSCs and effective peripheral lymphocyte deletion while preserving myeloid cells and regulatory T cells. Although Bcl-2 inhibition alone was insufficient to induce hematopoietic chimerism, the addition of a Bcl-2 inhibitor resulted in promotion of hematopoietic chimerism and renal allograft tolerance despite using only half of the dose of total body irradiation previously required. Selective inhibition of Bcl-2 is therefore a promising approach to induce hematopoietic chimerism without myelosuppression and has the potential to render HSCT more feasible for a variety of clinical indications.

Long-term Kinetics of Intragraft Gene Signatures in Renal Allograft Tolerance Induced by Transient Mixed Chimerism.

BACKGROUND: Renal allograft tolerance (TOL) has been successfully induced in nonhuman primates (NHPs) and humans through the induction of transient mixed chimerism. To elucidate the mechanisms of TOL, we compared local immunologic responses in renal allografts with those in T-cell-mediated rejection (TCMR) and chronic antibody-mediated rejection (CAMR) in NHPs. METHODS: Using the NanoString nCounter platform, we retrospectively studied 52 mRNAs in 256 kidney allograft samples taken from NHP kidney recipients of donor BMT. No immunosuppression was given after 1-month post-donor BMT. Recipients who achieved TOL (n = 13) survived for >1840 ± 1724 days with normal kidney function, while recipients with CAMR (n = 13) survived for 899 ± 550 days with compromised graft function, and recipients with TCMR (n = 15) achieved only short-term survival (132 ± 69 days). RESULTS: The most prominent difference between the groups was FOXP3, which was significantly higher in TOL than in CAMR and TCMR, both early (<1 y, P < 0.01) and late (≥1 y, P < 0.05) after transplant. Other mRNAs related to regulatory T cells (Treg), such as IL10, TGFB, and GATA3, were also high in TOL. In contrast, transcripts of inflammatory cytokines were higher in TCMR, while activated endothelium-associated transcripts were higher in CAMR than in TOL. The receiver operating characteristic analyses revealed that intragraft FOXP3 and CAV1 can reliably distinguish TOL from CAMR. CONCLUSIONS: High FOXP3 and other Treg-related mRNAs together with suppressed inflammatory responses and endothelial activation in renal allografts suggest that intragraft enrichment of Treg is a critical mechanism of renal allograft TOL induced by transient mixed chimerism.

Haploidentical hematopoietic cell and kidney transplantation for hematological malignancies and end-stage renal failure.

At Massachusetts General Hospital, we pioneered simultaneous hematopoietic cell (HCT)/kidney transplantation from HLA-identical related donors for the treatment of hematological malignancies with end-stage renal failure. We have now extended this to HLA-haploidentical donors in a pilot trial. Six recipients, 5 of whom were conditioned with fludarabine, cyclophosphamide, and total-body irradiation, underwent combined HCT/kidney transplantation from haploidentical donors; graft-versus-host disease (GVHD) prophylaxis included post-HCT cyclophosphamide, tacrolimus, and mycophenolate mofetil. One patient died as a result of complications of fludarabine neurological toxicity. No neurological toxicity was observed in subsequent patients who received lower fludarabine doses and more intense postfludarabine dialysis. There were no cases of grade 2 to 4 acute GVHD and 1 case of moderate chronic GVHD by 12 months. One patient experienced relapse of multiple myeloma at 30 months after HCT and died 4 years posttransplantation. Overall, 4 of 6 patients remain alive, without disease relapse and with long-term renal rejection-free survival. This trial was registered at www.clinicaltrials.gov as #NCT01758042.

Twenty-year Follow-up of Histocompatibility Leukocyte Antigen-matched Kidney and Bone Marrow Cotransplantation for Multiple Myeloma With End-stage Renal Disease: Lessons Learned.

BACKGROUND: Specific immune tolerance of transplanted organs in association with either transient or sustained lymphohematopoietic chimerism has been demonstrated in several preclinical animal models and clinically in patients who are full donor chimeras after hematopoietic stem cell transplantation and subsequently received kidney transplants from the same donor. Most recently, tolerance induction has been extended to patients in whom chimerism was intentionally induced at the time of kidney transplantation. METHODS: Twenty years ago, we reported the first successful histocompatibility leukocyte antigen-matched sibling donor bone marrow and kidney transplant following nonmyeloablative conditioning in a patient with multiple myeloma and end-stage renal disease (ESRD). After 2 decades, she has normal renal function in the absence of ongoing systemic immunosuppressive therapy. Nine patients have subsequently undergone similar treatment for multiple myeloma with ESRD. RESULTS: In the initial patient, hematopoietic chimerism was detectable for only 105 days after the transplant. In subsequent patients, chimerism detection ranged from 49 days to >14 years. Nevertheless, a long remission of the myeloma and long-term immunosuppression-free survival of the kidney allograft were achieved in 7 of the 10 patients, 5 of whom currently survive. CONCLUSIONS: This initial patient demonstrated the feasibility of performing combined histocompatibility leukocyte antigen-matched, sibling donor bone marrow and kidney transplantation for ESRD due to multiple myeloma. This experience paved the way for extending the initial trial to 9 additional patients with multiple myeloma and ESRD and, more recently, to tolerance induction strategies involving combined bone marrow and kidney transplantation for patients with and without an underlying malignancy.

Importance of Hematopoietic Mixed Chimerism for Induction of Renal Allograft Tolerance in Nonhuman Primates.

BACKGROUND: Although induction of durable mixed chimerism is required for murine skin allograft tolerance (TOL), renal allograft TOL has been achieved after induction of only transient mixed chimerism in nonhuman primates (NHPs) and humans. To better define the level/duration of chimerism required for stable renal allograft TOL, we retrospectively analyzed these parameters and compared them with transplant outcomes in NHP combined kidney and bone marrow transplant recipients. METHODS: Peripheral blood levels and duration of myeloid or lymphoid chimerism were retrospectively analyzed in 34 NHP combined kidney and bone marrow transplantation recipients which were divided into 3 groups: TOL, n = 10; chronic antibody-mediated rejection (CAMR), n = 12; and T cell-mediated rejection (TCMR), n = 12. RESULTS: All 4 of the recipients that failed to develop any chimerism lost their allografts due to TCMR after discontinuation of immunosuppression (56 ± 3 d). Among 30 recipients who successfully developed multilineage chimerism, 10 achieved long-term immunosuppression-free survival without rejection (1258 ± 388 d), 12 eventually developed CAMR (932 ± 155 d), and 8 developed TCMR (82 ± 10 d). The maximum level but not duration of lymphoid chimerism was significantly higher in TOL recipients compared with both CAMR (P = 0.0159) and TCMR (P = 0.0074). On the other hand, the maximum myeloid chimerism was significantly higher in TOL than in TCMR (P = 0.0469), but not in CAMR. Receiver operating characteristic analyses revealed that lymphoid chimerism levels of 3.1% or greater could reliably predict long-term immunosuppression-free renal allograft survival (P < 0.0001). CONCLUSIONS: This retrospective study confirmed that induction of chimerism is essential for long-term immunosuppression-free survival, which best correlates with lymphoid chimerism levels higher than 3.1%.

Long-term Nonhuman Primate Renal Allograft Survival Without Ongoing Immunosuppression in Recipients of Delayed Donor Bone Marrow Transplantation.

BACKGROUND: We have previously reported successful induction of renal allograft tolerance in nonhuman primates (NHP) after an initial posttransplant period of conventional immunosuppression (delayed tolerance) using a nonmyeloablative conditioning regimen consisting of anti-CD154 and anti-CD8 mAbs plus equine antithymocyte globulin (Atgam) and donor bone marrow transplantation (DBMT). Because these reagents are not currently clinically available, the protocol was revised to be applicable to human recipients of deceased donor allografts. METHOD: Four cynomolgus monkeys received major histocompatibility complex-mismatched kidney allografts with conventional immunosuppression for 4 months. The recipients were then treated with a nonmyeloablative conditioning regimen consisting of thymoglobulin, belatacept, and DBMT. The results were compared with recipients treated with conditioning regimen consisting of Atgam and anti-CD154 mAb, with and without anti-CD8 mAb. RESULTS: In 4 consecutive NHP recipients treated with the modified conditioning regimen, homeostatic recovery of CD8 TEM was delayed until after day 20 and multilineage chimerism was successfully induced. Three of the 4 recipients achieved long-term allograft survival (>728, >540, >449 days) without ongoing maintenance immunosuppression. Posttransplant MLR showed loss of antidonor CD8 T cell and CD4 IFNγ responses with expansion of CD4FOXP3 regulatory T cells. However, the late development of donor-specific antibody in NHP recipients confirms the need for additional anti-B-cell depletion with agents, such as rituximab, as has been shown in our clinical trials. CONCLUSIONS: This study provides proof of principle that induction of mixed chimerism and long-term renal allograft survival without immunosuppression after delayed DBMT is possible with clinically available reagents.

Effect of Ex Vivo-Expanded Recipient Regulatory T Cells on Hematopoietic Chimerism and Kidney Allograft Tolerance Across MHC Barriers in Cynomolgus Macaques.

BACKGROUND: Infusion of recipient regulatory T (Treg) cells promotes durable mixed hematopoietic chimerism and allograft tolerance in mice receiving allogeneic bone marrow transplant (BMT) with minimal conditioning. We applied this strategy in a Cynomolgus macaque model. METHODS: CD4 CD25 Treg cells that were polyclonally expanded in culture were highly suppressive in vitro and maintained high expression of FoxP3. Eight monkeys underwent nonmyeloablative conditioning and major histocompatibility complex mismatched BMT with or without Treg cell infusion. Renal transplantation (from the same BMT donor) was performed 4 months post-BMT without immunosuppression to assess for robust donor-specific tolerance. RESULTS: Transient mixed chimerism, without significant T cell chimerism, was achieved in the animals that received BMT without Treg cells (N = 3). In contrast, 2 of 5 recipients of Treg cell BMT that were evaluable displayed chimerism in all lineages, including T cells, for up to 335 days post-BMT. Importantly, in the animal that survived long-term, greater than 90% of donor T cells were CD45RA CD31, suggesting they were new thymic emigrants. In this animal, the delayed (to 4 months) donor kidney graft was accepted more than 294 days without immunosuppression, whereas non-Treg cell BMT recipients rejected delayed donor kidneys within 3 to 4 weeks. Early CMV reactivation and treatment was associated with early failure of chimerism, regardless of Treg cell administration. CONCLUSIONS: Our studies provide proof-of-principle that, in the absence of early CMV reactivation (and BM-toxic antiviral therapy), cotransplantation of host Treg cell can promote prolonged and high levels of multilineage allogeneic chimerism and robust tolerance to the donor.

Induced regulatory T cells in allograft tolerance via transient mixed chimerism.

Successful induction of allograft tolerance has been achieved in nonhuman primates (NHPs) and humans via induction of transient hematopoietic chimerism. Since allograft tolerance was achieved in these recipients without durable chimerism, peripheral mechanisms are postulated to play a major role. Here, we report our studies of T cell immunity in NHP recipients that achieved long-term tolerance versus those that rejected the allograft (AR). All kidney, heart, and lung transplant recipients underwent simultaneous or delayed donor bone marrow transplantation (DBMT) following conditioning with a nonmyeloablative regimen. After DBMT, mixed lymphocyte culture with CFSE consistently revealed donor-specific loss of CD8+ T cell responses in tolerant (TOL) recipients, while marked CD4+ T cell proliferation in response to donor antigens was found to persist. Interestingly, a significant proportion of the proliferated CD4+ cells were FOXP3+ in TOL recipients, but not in AR or naive NHPs. In TOL recipients, CD4+FOXP3+ cell proliferation against donor antigens was greater than that observed against third-party antigens. Finally, the expanded Tregs appeared to be induced Tregs (iTregs) that were converted from non-Tregs. These data provide support for the hypothesis that specific induction of iTregs by donor antigens is key to long-term allograft tolerance induced by transient mixed chimerism.

Effect of tolerance versus chronic immunosuppression protocols on the quality of life of kidney transplant recipients.

BACKGROUND: Kidney transplant patients on tolerance protocols avoid the morbidity associated with the use of conventional chronic immunosuppressive regimens. However, the impact of tolerance versus conventional regimens on the quality of life (QOL) of kidney transplant patients is unknown. METHODS: Five patients who achieved long-term immunosuppression-free renal allograft survival after combined kidney and bone marrow transplantation (tolerant group) were compared with thirty-two comparable kidney transplant recipients on conventional immunosuppression (conventional group). QOL was compared with 16 conventional recipients using the Kidney Disease Quality of Life Short Form 36 (KDQOL SF-36) and the Modified Transplant Symptom Occurrence and Symptom Distress Scale (MTSOSD-59R). RESULTS: Patients in the tolerant group required significantly less treatment after transplant for hypertension and no medications for diabetes (P < 0.01). There was no incidence of diabetes, dyslipidemia, or malignancies in the tolerant group, while these were observed in 12.5%, 40.6%, and 11.8% of the conventional group, respectively. Tolerant patients experienced better overall health (P < 0.01) and scored higher on kidney transplant-targeted scales and healthy survey scales than patients in the conventional group according to the KDQOL SF-36 (P < 0.05). Tolerant patients were less likely to experience depression, dyspnea, excessive appetite/thirst, flatulence, hearing loss, itching, joint pain, lack of energy, muscle cramps, and lack of libido than conventional patients according to the MTSOSD-59R (P < 0.05). CONCLUSION: Kidney transplant recipients who achieved tolerance experience significantly fewer incidences of complications, improved QOL, and fewer comorbid symptoms compared with patients on conventional immunosuppression. These results support the expanded use of tolerance protocols in kidney transplantation.

Clinical trials for induction of renal allograft tolerance.

PURPOSE OF REVIEW: This review summarizes the current state of clinical trials for tolerance induction of human leukocyte antigen-matched or mismatched renal allografts via peritransplant infusion of donor bone marrow-derived products. Recent efforts to apply infusion of expanded regulatory T-cell preparations to minimize immunosuppressive dosages are also reviewed. RECENT FINDINGS: Three centers in the United States have reported clinical trials for tolerance induction in recipients of living donor kidney transplants via donor hematopoietic stem cell transplantation. They have observed varying degrees of successful renal allograft tolerance induction following the establishment of either transient or persistent donor chimerism.A more recent clinical trial planned to evaluate administration of regulatory T cells to harness the immune response has recently been initiated in eight centers in Europe and the United States. SUMMARY: Tolerance induction in clinical kidney transplantation from live donors has been achieved by donor hematopoietic stem cell transplantation. Improving the consistency and safety of tolerance induction and extending successful protocols to other organs, as well as to organs from deceased donors, are critical next steps to bringing tolerance to a wider range of clinical applications.

Clinical significance of microscopic melanoma metastases in the nonhottest sentinel lymph nodes.

IMPORTANCE: A practice gap exists in the surgical removal of sentinel lymph nodes, from removal of only the most radioactive (hottest) lymph node to removal of all lymph nodes with radioactivity greater than 10% of the hottest lymph node. OBJECTIVE: To determine the clinical significance of melanoma in sentinel lymph nodes that are not the hottest sentinel node and to determine the risk for disease progression based on sentinel lymph node status and primary tumor characteristics. DESIGN, SETTING, AND PARTICIPANTS: Consecutive patients with cutaneous melanoma with sentinel lymph nodes resected from January 5, 2004, to June 30, 2008, with a mean follow-up of 59 months, at Massachusetts General Hospital were included in this retrospective review. The last year of follow-up was 2012. The operative protocol led to resection of all sentinel lymph nodes with radioactivity greater than 10% of the hottest lymph node. The number of lymph nodes removed, technetium-99m counts for each sentinel lymph node, presence or absence of sentinel lymph node metastases, primary tumor characteristics, disease progression, and melanoma-specific survival were recorded. MAIN OUTCOMES AND MEASURES: Microscopic melanoma metastases in the hottest and nonhottest sentinel lymph nodes and factors that correlate with disease progression and mortality. RESULTS: A total of 1575 sentinel lymph nodes were analyzed in 475 patients. Ninety-one patients (19%) had positive sentinel lymph nodes. Of these, 72 (79%) had metastases in the hottest sentinel lymph node. Of 19 cases with tumor present, but not in the hottest sentinel lymph node, counts ranged from 26% to 97% of the hottest node. Progression occurred in 43% of patients with sentinel node metastasis, regardless of whether the hottest lymph node was positive. In patients with negative sentinel lymph nodes, 11% developed metastases beyond the sentinel lymph node basin and 3.4% recurred in the basin. Mitogenicity of the primary tumor was associated with mortality (odds ratio, 2.435; 95% CI, 1.351-4.391; P < .001). Removing only the hottest sentinel lymph node would have led to false-negative results in 19 of 475 (4%) of all patients and 19 of 91 patients (21%) with positive sentinel lymph nodes. The 8-year survival in patients with at least 1 positive sentinel lymph node was less than 55%. The presence of more than 1 mitosis per square millimeter in the primary cutaneous melanoma was associated with decreased survival. CONCLUSIONS AND RELEVANCE: Microscopic melanoma metastases was associated with disease progression and mortality, whether present in the hottest sentinel lymph node or not. These observations emphasize the importance of removing the less hot nodes, addressing a practice gap in the surgical approach to patients with melanoma.

Outcome of patients with de novo versus nevus-associated melanoma.

BACKGROUND: Prior reports indicate a wide range of melanomas in histopathologic contiguity with a nevus, and an associated nevus has unclear prognostic implications in melanoma. OBJECTIVE: We sought to investigate the relationship among nevus-associated melanomas, sentinel lymph node status, and overall survival. METHODS: We conducted a retrospective analysis of 850 patients with cutaneous melanoma and sentinel lymph node removed at Massachusetts General Hospital from 1998 through 2008 and meta-analysis of the literature. RESULTS: Nevus-associated melanomas represented 28% (235/850) of cases and were significantly correlated with younger age (P = .03), truncal site (P = .0005), superficial spreading type (P < .0001), and absent ulceration (P = .005). There was no association with sentinel lymph node status (P = .94) and no survival difference between nevus-associated versus de novo melanoma (P = .41). Meta-analysis of over 4000 cases revealed a similar percentage of associated nevi (32%). LIMITATIONS: This was a retrospective study. CONCLUSIONS: Approximately 30% of melanomas are associated with a nevus. The presence of a nevus associated with a melanoma has no prognostic implication in sentinel lymph node status or overall survival.

Clinical strategy for induction of transplantation tolerance through mixed chimerism.

Following the demonstration that transplant tolerance could be induced in non-human primate recipients treated with non-myeloablative conditioning that resulted in only transient chimerism, we began in 1998 to evaluate this approach first in patients with end stage renal disease (ESRD), secondary to multiple myeloma (MM). A total of 10 patients with ESRD and MM have been treated with this initial protocol. Only 2 recipients developed evidence of reversible renal allograft rejection after stopping immunosuppression. Long-term (up to 14 years) operational renal allograft tolerance has been observed in all 10 patients, even in those with transient hematopoietic chimerism. Control of the MM has been less complete, as recurrent disease developed in five of these patients, three of whom expired. Nevertheless, in view of the essentially 100% 3-5 year mortality typically expected with alternative treatments for this challenging population, it has been suggested that combined kidney and donor bone marrow transplantation (CKBMT) following non-myeloablative conditioning should become the standard therapy for patients with ESRD secondary to MM (27). These encouraging results, as well as the acceptable morbidity observed in cancer patients receiving non-myeloablative human leukocyte antigen (HLA)-mismatched bone marrow transplantation in our center, led us to next evaluate CKBMT in 10 patients of HLA-mismatched transplants. All 10 subjects developed transient chimerism and in seven of these, immunosuppression was successfully discontinued. Four subjects continue to be immunosuppression free for periods of 4.5-11 years, while in three, reinstitution of immunosuppression was advised or accomplished after 5-8 years due to recurrence of original disease or chronic antibody mediated rejection. Donor-specific antibodies were frequently detectable in the earlier recipients. In contrast, no donor-specific antibodies were detected after immunosuppression was discontinued in the last four subjects, who showed more consistent and durable B-cell depletion. In conclusion, our studies have shown that immunosuppression-free renal allograft survival can be achieved in both HLAmatched and mismatched donor-recipient pairs, with follow up times now of greater than 14 years after induction of transient chimerism through CKBMT.

Urinary reconstruction after kidney transplantation: pyeloureterostomy or ureteroneocystostomy.

PURPOSE: Ureteroneocystostomy (UCN) is the most widely used urinary reconstruction technique during kidney transplantation. Disadvantages of this technique include a high incidence of hematuria and reflux, plus the potential for obstruction resulting from distal ureteral fibrosis. Pyeloureterostomy (PU) avoids these complications but increases the technical complexity. METHODS: Between January 1990 and December 2005, 1066 adults patients underwent kidney transplantations; 768 patients (72.1%) had urinary reconstruction by PU and 298 (27.9%) underwent UNC. RESULTS: Patients in the PU group underwent simultaneous ipsilateral native nephrectomy. The operative time was longer in the PU group compared with the UNC group: 210 ± 36 min versus 182 ± 24 min (P < 0.001). Overall surgical complications in the PU group were comparable to those in the UNC group (9.5% versus 12.3%). The urinary complication rate was also comparable in both groups: 3.2% (25 of 768) in the PU group and 5% (15 of 298) in the UNC group. However, urinary obstruction comprised 60% of urinary complications in the UNC group, compared with 32% in the PU group (P < 0.01). We treated most urinary complications non-operatively. However, 24% of patients (six of 25) in the PU group needed operative intervention or revision for ureteral reconstruction, compared with 46.6% (seven of 15) in the UNC group (P < 0.01). CONCLUSIONS: Pyeloureterostomy is a safe and effective method for urinary tract reconstruction in renal transplantation. Pyeloureterostomy should be part of every transplant surgeon's armamentarium.

The distribution of microscopic melanoma metastases in sentinel lymph nodes: implications for pathology protocols.

The utility of sectioning at multiple levels in the histopathologic analysis of sentinel lymph nodes (SLNs) for melanoma and the correlation of metastasis size with risk of subsequent metastasis were investigated. Metastatic melanoma was identified in SLNs from 91 of 475 (19%) melanoma patients with SLN sampling at the Massachusetts General Hospital between 2004 and 2008. All SLNs were evaluated by a 9-slide protocol: sets of MART-1, hematoxylin and eosin, and S100 stains at 3 distinct levels separated by 80 µm. The location and size of the tumor deposits were evaluated in the context of subsequent metastasis and overall survival. Of the 91 patients with positive sentinel nodes, all 9 protocol slides were available for review in 61 (67%). Eleven of 61 patients had no tumor present in the first set of levels; 2 of these patients died of metastatic melanoma. Patients in whom 11 or more tumor cells were detected in the sentinel node had a greater chance of developing subsequent metastases when compared with patients in whom 10 or fewer tumor cells were detected (P=0.05). Of those with either metastases >2 mm in diameter or extracapsular extension, 50% developed metastases beyond the SLN basin. Eliminating 1 of the 3 levels in the SLN detection protocol would have led to a false-negative diagnosis in 18% of patients.

Antithymocyte globulin induction in living donor renal transplant recipients: final report of the TAILOR registry.

BACKGROUND: The Thymoglobulin Antibody Immunosuppression in Living Donor Recipients registry was established to assess clinical experience with rabbit antithymocyte globulin (rATG; Thymoglobulin) in living donor renal transplant recipients. METHODS: From 2003 to 2008, US transplant centers prospectively entered information on patients who received rATG induction. In addition to standard United Network for Organ Sharing registry data elements, information was collected regarding immunosuppression, viral prophylaxis, acute rejection, and adverse events. RESULTS: Data on 2322 patients from 49 transplant centers were enrolled and met inclusion criteria for analysis. Patient and graft survival were 99.3% and 99.0% at 6 months and 98.4% and 98.2% at 12 months as recorded in Thymoglobulin Antibody Immunosuppression in Living Donor Recipients registry and were 91.5% and 83.2% at 5 years by Kaplan-Meier estimates based on linked United Network for Organ Sharing registry records. Freedom from rejection was 93.6% through 5 years. Mean rATG cumulative dose was 5.29 mg/kg. More than one-third of patients (37.6%) were steroid-free at discharge, and nearly half of patients (48%) were steroid-free at 12 months. Before discharge, 3.2% experienced serious adverse events, with 11 events (0.005%) reported as possibly or probably related to rATG. Incidence of cytomegalovirus infection was 4.2% at 12 months, and 99.1% of patients were posttransplant lymphoproliferative disorder-free through 5 years. CONCLUSIONS: rATG induction in living donor renal transplantation is safe and associated with a low incidence of acute rejection and posttransplantation complications.

Differential effects of denileukin diftitox IL-2 immunotoxin on NK and regulatory T cells in nonhuman primates.

Denileukin diftitox (DD), a fusion protein comprising IL-2 and diphtheria toxin, was initially expected to enhance antitumor immunity by selectively eliminating regulatory T cells (Tregs) displaying the high-affinity IL-2R (α-ß-γ trimers). Although DD was shown to deplete some Tregs in primates, its effects on NK cells (CD16(+)CD8(+)NKG2A(+)CD3(-)), which constitutively express the intermediate-affinity IL-2R (ß-γ dimers) and play a critical role in antitumor immunity, are still unknown. To address this question, cynomolgus monkeys were injected i.v. with two doses of DD (8 or 18 µg/kg). This treatment resulted in a rapid, but short-term, reduction in detectable peripheral blood resting Tregs (CD4(+)CD45RA(+)Foxp3(+)) and a transient increase in the number of activated Tregs (CD4(+)CD45RA(-)Foxp3(high)), followed by their partial depletion (50-60%). In contrast, all NK cells were deleted immediately and durably after DD administration. This difference was not due to a higher binding or internalization of DD by NK cells compared with Tregs. Coadministration of DD with IL-15, which binds to IL-2Rß-γ, abrogated DD-induced NK cell deletion in vitro and in vivo, whereas it did not affect Treg elimination. Taken together, these results show that DD exerts a potent cytotoxic effect on NK cells, a phenomenon that might impair its antitumoral properties. However, coadministration of IL-15 with DD could alleviate this problem by selectively protecting potentially oncolytic NK cells, while allowing the depletion of immunosuppressive Tregs in cancer patients.

Liver transplantation results in complete neurologic recovery from malignant hypertension secondary to fulminant hepatic failure: a case report.

BACKGROUND: Uncontrolled intracranial hypertension can lead to cerebral herniation and death in patients with acute liver failure. CASE REPORT: A 26-year-old female was admitted for acute liver failure following inadvertent acetaminophen overdose. The pH on admission was 6.9. Her neurologic status precipitously deteriorated and she was listed for liver transplantation. An intracranial pressure (ICP) monitoring catheter was inserted, which revealed a pressure >60 mmHg. After neurointensive care treatment, ICP was lowered and an emergency left lobe living donor liver transplant was performed. Intraoperative management of the ICP, which rose to 80 mmHg during the explant phase, was achieved by therapy with barbiturates and hypothermia. After surgery, hepatic function improved initially, but 7 days post transplantation the graft showed signs of acute failure. The pathology report of a liver biopsy suggested acute rejection and liver retransplantation using a deceased donor liver was then carried out. The postoperative course was uneventful and the patient recovered completely without any residual neurologic deficits. CONCLUSIONS: This case states that favourable outcomes can result from sub-optimal starting points, and that the human brain has the ability to overcome extremely adverse conditions. Critical in this effort is the role of proper neuromonitoring which helps implement the appropriate treatment measures.

Contributions of direct and indirect alloresponses to chronic rejection of kidney allografts in nonhuman primates.

The relative contribution of direct and indirect allorecognition pathways to chronic rejection of allogeneic organ transplants in primates remains unclear. In this study, we evaluated T and B cell alloresponses in cynomolgus monkeys that had received combined kidney/bone marrow allografts and myeloablative immunosuppressive treatments. We measured donor-specific direct and indirect T cell responses and alloantibody production in monkeys (n = 5) that did not reject their transplant acutely but developed chronic humoral rejection (CHR) and in tolerant recipients (n = 4) that never displayed signs of CHR. All CHR recipients exhibited high levels of anti-donor Abs and mounted potent direct T cell alloresponses in vitro. Such direct alloreactivity could be detected for more than 1 y after transplantation. In contrast, only two of five monkeys with CHR had a detectable indirect alloresponse. No indirect alloresponse by T cells and no alloantibody responses were found in any of the tolerant monkeys. Only one of four tolerant monkeys displayed a direct T cell alloresponse. These observations indicate that direct T cell alloresponses can be sustained for prolonged periods posttransplantation and result in alloantibody production and chronic rejection of kidney transplants, even in the absence of detectable indirect alloreactivity.