Faculty Bibliography

BACKGROUND:Xenografts from genetically modified pigs have become one of the most promising solutions to the dearth of human organs available for transplantation. The challenge in this model has been hyperacute rejection. To avoid this, pigs have been bred with a knockout of the alpha-1,3-galactosyltransferase gene and with subcapsular autologous thymic tissue. METHODS:We transplanted kidneys from these genetically modified pigs into two brain-dead human recipients whose circulatory and respiratory activity was maintained on ventilators for the duration of the study. We performed serial biopsies and monitored the urine output and kinetic estimated glomerular filtration rate (eGFR) to assess renal function and xenograft rejection. RESULTS:in Recipient 2. In both recipients, the creatinine level, which had been at a steady state, decreased after implantation of the xenograft, from 1.97 to 0.82 mg per deciliter in Recipient 1 and from 1.10 to 0.57 mg per deciliter in Recipient 2. The transplanted kidneys remained pink and well-perfused, continuing to make urine throughout the study. Biopsies that were performed at 6, 24, 48, and 54 hours revealed no signs of hyperacute or antibody-mediated rejection. Hourly urine output with the xenograft was more than double the output with the native kidneys. CONCLUSIONS:Genetically modified kidney xenografts from pigs remained viable and functioning in brain-dead human recipients for 54 hours, without signs of hyperacute rejection. (Funded by Lung Biotechnology.).

BACKGROUND:Immunotherapy, specifically immune checkpoint inhibitors (ICIs), including anti-programmed cell death 1 (anti-PD1), has recently received clinical approval for the treatment of adult hepatocellular carcinoma (HCC). However, the safety and efficacy of ICIs prior to solid organ transplant are unknown, especially in pediatrics. Safety reports are variable in adults, with some series describing subsequent allograft rejection and loss while others report successful transplants without allograft rejection.As ICIs stimulate the immune system by blocking the interaction between PD1 and the ligand-receptor pair programmed cell death-ligand 1 (PDL1), the downstream effects of T-cell activation increase the risk of graft rejection. METHODS:Here, we present a case of an adolescent with moderately differentiated non-fibrolamellar HCC treated with pembrolizumab, an anti-PD1 therapy, who subsequently underwent successful orthotopic liver transplantation (OLT). RESULTS:Our patient received an OLT 138 days from the last pembrolizumab dose with graft preservation. The patient has no evidence of recurrent disease or any episode of allograft rejection 48 months post OLT. Staining of tumor and normal tissues from longitudinal specimens finds PDL1 positive Kupffer cells present in normal liver and peritumoral areas with no changes post anti-PD1 therapy. In contrast, tumor cells were negative for PDL1. CONCLUSION/CONCLUSIONS:This case represents a basis for optimism in potential use of anti-PD1 therapy in liver transplant candidates and supports further investigation of immune checkpoint inhibitors use in this unique patient population.

BACKGROUND:Hepatic resections are uncommon in children. Most studies reporting complications of these procedures and risk factors associated with unplanned readmissions are limited to retrospective data from single centers. We investigated risk factors for 30-day unplanned readmission after hepatectomy in children using the American College of Surgeons National Surgical Quality Improvement-Pediatric database. METHODS:The database was queried for patients aged 0-18 years who underwent hepatectomy for the treatment of liver lesions from 2012 to 2018. Chi-squared tests were performed to evaluate for potential risk factors for unplanned readmissions. A multivariate regression analysis was performed to identify independent predictors for unplanned 30-day readmissions. RESULTS:Among 438 children undergoing hepatectomy, 64 (14.6%) had unplanned readmissions. The median age of the hepatectomy cohort was 1 year (0-17); 55.5% were male. Patients readmitted had significantly higher rates of esophageal/gastric/intestinal disease (26.56% vs. 14.97%; p=0.022), current cancer (85.94% vs. 75.67%; p=0.012), and enteral and parenteral nutritional support (31.25% vs. 17.65%; p=0.011). Readmitted patients had significantly higher rates of perioperative blood transfusion (67.19% vs. 52.41%; p=0.028), organ/space surgical site infection (10.94% vs. 1.07%; p<.001), sepsis (15.63% vs. 3.74%; p<.001), and total parenteral nutrition at discharge (9.09% vs. 2.66%; p=0.041). Organ/space surgical site infection was an independent risk factor for unplanned readmission (OR=9.598, CI [2.070-44.513], p=0.004) by multivariable analysis. CONCLUSION:Unplanned readmissions after liver resection are frequent in pediatric patients. Organ/space surgical site infections may identify patients at increased risk for unplanned readmission. Strategies to reduce these complications may decrease morbidity and costs associated with unplanned readmissions.

In humans receiving intestinal transplantation (ITx), long-term multilineage blood chimerism often develops. Donor T cell macrochimerism (≥4%) frequently occurs without graft-versus-host disease (GVHD) and is associated with reduced rejection. Here we demonstrate that patients with macrochimerism had high graft-versus-host (GvH) to host-versus-graft (HvG) T cell clonal ratios in their allografts. These GvH clones entered the circulation, where their peak levels were associated with declines in HvG clones early after transplant, suggesting that GvH reactions may contribute to chimerism and control HvG responses without causing GVHD. Consistently, donor-derived T cells, including GvH clones, and CD34+ hematopoietic stem and progenitor cells (HSPCs) were simultaneously detected in the recipients' BM more than 100 days after transplant. Individual GvH clones appeared in ileal mucosa or PBMCs before detection in recipient BM, consistent with an intestinal mucosal origin, where donor GvH-reactive T cells expanded early upon entry of recipient APCs into the graft. These results, combined with cytotoxic single-cell transcriptional profiles of donor T cells in recipient BM, suggest that tissue-resident GvH-reactive donor T cells migrated into the recipient circulation and BM, where they destroyed recipient hematopoietic cells through cytolytic effector functions and promoted engraftment of graft-derived HSPCs that maintain chimerism. These mechanisms suggest an approach to achieving intestinal allograft tolerance.

The therapeutic applications of regulatory T cells (T_regs ) include treating autoimmune diseases, graft-versus-host disease and induction of transplantation tolerance. For ex-vivo expanded T_regs to be used in deceased donor transplantation, they must be able to suppress T cell responses to a broad range of human leukocyte antigen (HLA). Here, we present a novel approach for the expansion of polyspecific T_regs in cynomolgus macaques that was adapted from a good manufacturing practice-compliant protocol. T_regs were isolated by fluorescence-activated cell sorting (FACS) and expanded in the presence of a panel of CD40L-stimulated B cells (CD40L-sBc). Prior to T_reg culture, CD40L-sBc were expanded in vitro from multiple major histocompatibility complex (MHC)-disparate macaques. Expanded T_regs expressed high levels of forkhead box protein 3 (FoxP3) and Helios, a high percentage of T_reg -specific demethylated region (TSDR) demethylation and strong suppression of naïve T cell responses in vitro. In addition, these T_regs produced low levels of inflammatory cytokines and were able to expand post-cryopreservation. Specificity assays confirmed that these T_regs were suppressive upon activation by any antigen-presenting cells (APCs) whose MHC was shared by CD40L-sBc used during expansion, proving that they are polyspecific. We developed an approach for the expansion of highly suppressive cynomolgus macaque polyspecific T_regs through the use of a combination of CD40L-engineered B cells with the potential to be translated to clinical studies. To our knowledge, this is the first report that uses a pool of MHC-mismatched CD40L-sBc to create polyspecific T_regs suitable for use in deceased-donor transplants.