RNA interference therapeutics in organ transplantation:The dawn of a new era

RNA interference (RNAi) is a natural process through which double-stranded RNA molecules can silence the gene carrying the same code as the particular RNA of interest. In 2006, the discovery of RNAi was awarded the Nobel Prize in Medicine and its success has accumulated since. Gene silencing through RNAi has been used successfully in a broad range of diseases, and, more recently, this technique has gained interest in the field of organ transplantation. Here, genes related to ischemia-reperfusion injury (IRI) or graft rejection may be silenced to improve organ quality after transplantation. Several strategies have been used to deliver siRNA, and pretransplant machine perfusion presents a unique opportunity to deliver siRNA to the target organ during ex situ preservation. In this review, the potential of RNAi in the field of organ transplantation will be discussed. A brief overview on the discovery of RNAi, its mechanism, and limitations are included. In addition, studies using RNAi to target genes related to IRI in liver, kidney, lung, and heart transplantation are discussed

The Emerging Role of Viability Testing During Liver Machine Perfusion

The transplant community continues to be challenged by the disparity between the need for liver transplantation and the shortage of suitable donor organs. At the same time, the number of unused donor livers continues to increase, most likely attributed to the worsening quality of these organs. To date, there is no reliable marker of liver graft viability that can predict good posttransplant outcomes. Ex situ machine perfusion offers additional data to assess the viability of donor livers before transplantation. Hence, livers initially considered unsuitable for transplantation can be assessed during machine perfusion in terms of appearance and consistency, hemodynamics, and metabolic and excretory function. In addition, postoperative complications such as primary nonfunction or posttransplant cholangiopathy may be predicted and avoided. A variety of viability criteria have been used in machine perfusion, and to date there is no widely accepted composition of criteria for clinical use. This review discusses potential viability markers for hepatobiliary function during machine perfusion, describes current limitations, and provides future recommendations for the use of viability criteria in clinical liver transplantation

Donor Diabetes and Prolonged Cold Ischemia Time Synergistically Increase the Risk of Graft Failure After Liver Transplantation

BACKGROUND: Both prolonged cold ischemia time (CIT) and donor history of diabetes mellitus (DM) are associated with reduced graft survival after liver transplantation. However, it is unknown whether the adverse effect of prolonged CIT on posttransplant graft survival is more pronounced after transplant with DM versus non-DM donor grafts. METHODS: The study sample included 58 226 liver transplant recipients (2002-2015) from the Scientific Registry of Transplant Recipients. Multivariable Cox survival regression with interaction analysis was used to quantify the extent to which history of donor DM (n = 6478) potentiates the adverse effect of prolonged ( \u3e /=8 hours) CIT (n = 18 287) on graft survival. RESULTS: Donor DM and CIT 8 hours or longer were each associated with increased risk of graft failure (GF) (adjusted hazard ratio [aHR], 1.19; 95% confidence interval [CI], 1.06-1.35 and aHR, 1.42; 95% CI, 1.32-1.53, respectively) compared with transplanted grafts without either risk factor. However, the combination of DM and CIT 8 hours or longer was associated with a higher risk of GF than either factor alone (aHR, 1.79; 95% CI, 1.55-2.06) and had a synergy index of 1.30. The interaction was significant on a multiplicative scale in the later postoperative period, days 31 to 365 (P = 0.047). CONCLUSIONS: These results suggest that liver grafts from DM donors are more susceptible to the adverse effects of prolonged CIT than livers from non-DM donors. We need to be cognizant that they are more susceptible to ischemic injury, and this may be considered during the allocation process

Heat Stroke as a Cause of Liver Failure and Evaluation of Liver Transplant

Heat stroke is a multiple organ dysfunction syndrome of poorly understood pathogenesis. Exertional heat stroke with acute liver failure is a rarely reported condition. Liver transplant has been recommended as treatment in cases of severe liver dysfunction; however, there are only 5 described cases of long-term survival after this procedure in patients with heat stroke. Here, we present 2 cases of young athletes who developed heat stroke. Both patients developed acute liver failure and were listed for liver transplant. Liver function tests of one patient improved, and he was discharged on postoperative day 13. The other patient showed no signs of improvement and liver biopsy showed massive necrosis. The patient underwent combined kidney-liver transplant and was discharged on postoperative day 17. After a follow-up of longer than 6 years, both patients are doing well with normal liver function and no neurologic sequelae. We also reviewed all published cases of hepatic failure associated with heat stroke and found 9 published cases of liver transplant for heat stroke in the English literature. Conservative management appears to be justified in heat stroke-associated liver failure, even in the presence of accepted criteria for emergency liver transplant

Single-Center Experience With Liver Transplant Using Donors With Very High Transaminase Levels

OBJECTIVES: Elevation of transaminases has been used as a marker of hepatic ischemic injury and as a crucial parameter for liver graft assessment. However, analysis of serum transaminases has limitations regarding the quantitative assessment of liver necrosis and is not a reliable predictor of outcomes. MATERIALS AND METHODS: We retrospectively reviewed the medical records of all liver transplants (N = 238) performed at the UMass Memorial Medical Center from 2009 to 2013. RESULTS: Fourteen liver grafts showed high peak aminotransferases alanine aminotransferase (ALT) and aspartate aminotransferase (AST) at \u3e 1000 U/L. This high aminotransferase group was compared with 224 donors with low transaminase levels (ALT/AST \u3c 1000 U/L). The high transaminase donors had a median peak AST level of 3216 U/L (range, 1823-13?030 U/L) and ALT level of 2677 U/L (range, 812-7080 U/L). The high transaminase donors showed higher levels of lactate dehydrogenase, international normalized ratio, total bilirubin, and gamma-glutamyltransferase compared with low transaminase donors; however, only lactate dehydrogenase results reached statistical significance. None of the grafts from the high transaminase donors showed primary nonfunction. Three-year graft and patient survival rates were similar in both groups (75% vs 80% [P = .48] and 72% vs 82% [P = .33], respectively). In an analysis of the discard rate of livers over a 10-year period in the United States using the Scientific Registry of Transplant Recipients database, the discard rate of livers with high aminotransferase levels was 69.14% compared with 22.23% for livers with low transaminase levels. CONCLUSIONS: Liver grafts from donors with high transaminase levels can lead to clinical results that are similar to liver grafts from donors who had lower peak transaminase levels

A Comparative Study of Single and Dual Perfusion During End-ischemic Subnormothermic Liver Machine Preservation

Background: It remains controversial if arterial perfusion in addition to portal vein perfusion during machine preservation improves liver graft quality. Comparative studies using both techniques are lacking. We studied the impact of using single or dual machine perfusion of donation after circulatory death rat livers. In addition, we analyzed the effect of pulsatile versus continuous arterial flow. Methods: Donation after circulatory death rat livers (n = 18) were preserved by 6 hours cold storage, followed by 1 hour subnormothermic machine perfusion (20 degrees C, pressure of 40/5 mm Hg) and 2 hours ex vivo warm reperfusion (37 degrees C, pressure of 80/11 mm Hg, 9% whole blood). Machine preservation was either through single portal vein perfusion (SP), dual pulsatile (DPP), or dual continuous perfusion (DCP) of the portal vein and hepatic artery. Hydrodynamics, liver function tests, histopathology, and expression of endothelial specific genes were assessed during 2 hours warm reperfusion. Results: At the end of reperfusion, arterial flow in DPP livers tended to be higher compared to DCP and SP grafts. However, this difference was not significant nor was better flow associated with better outcome. No differences in bile production or alanine aminotransferase levels were observed. SP livers had significantly lower lactate compared to DCP, but not DPP livers. Levels of Caspase-3 and tumor necrosis factor-alpha were similar between the groups. Expression of endothelial genes Kruppel-like-factor 2 and endothelial nitric oxide synthase tended to be higher in dual perfused livers, but no histological evidence of better preservation of the biliary endothelium or vasculature of the hepatic artery was observed. Conclusions: This study shows comparable outcomes after using a dual or single perfusion approach during end-ischemic subnormothermic liver machine preservation

Is single portal vein perfusion the best approach for machine preservation of liver grafts

We have read with great interest the article by Schlegel et al., describing a single portal vein approach for hypothermic oxygenated machine perfusion (HOPE) of donation after circulatory death (DCD) liver grafts. It is unquestionable that single portal vein perfusion adds simplicity to liver machine perfusion and may provide protection. However, the main question that needs to be addressed is whether this is the best perfusion technique

Gene Silencing With siRNA (RNA Interference): A New Therapeutic Option During Ex Vivo Machine Liver Perfusion Preservation

RNA interference (RNAi) is a natural process of posttranscriptional gene regulation that has raised a lot of attention culminating with the Nobel Prize in Medicine in 2006. RNAi-based therapeutics have been tested in experimental transplantation to reduce ischemia/reperfusion injury (IRI) with success. Modulation of genes of the innate immune system, as well as apoptotic genes, and those involved in the nuclear factor kappa B pathways can reduce liver injury in rodent liver pedicle clamping and transplantation models of IRI. However, in vivo use of RNAi faces limitations regarding the method of administration, uptake, selectivity, and stability. Machine perfusion preservation, a more recent alternative approach for liver preservation showing superior results to static cold preservation, could be used as a platform for gene interference therapeutics. Our group was the first to demonstrate uptake of small interfering RNA (siRNA) during liver machine preservation under both normothermic and hypothermic perfusion. Administering siRNA in the perfusion solution during ex vivo machine preservation has several advantages, including more efficient delivery, lower doses and cost-saving, and none/fewer side effects to other organs. Recently, the first RNAi drug was approved by the US Food and Drug Administration for clinical use, opening a new avenue for new drugs with different clinical applications. RNAi has the potential to have transformational therapeutic applications in several areas of medicine including transplantation. We believe that machine preservation offers great potential to be the ideal delivery method of siRNA to the liver graft, and future studies should be initiated to improve the clinical applicability of RNAi in solid organ transplantation

Prolonged hypothermic machine perfusion enables daytime liver transplantation - an IDEAL stage 2 prospective clinical trial

Background: Liver transplantation is traditionally performed around the clock to minimize organ ischemic time. However, the prospect of prolonging preservation times holds the potential to streamline logistics and transform liver transplantation into a semi-elective procedure, reducing the need for nighttime surgeries. Dual hypothermic oxygenated machine perfusion (DHOPE) of donor livers for 1–2 h mitigates ischemia-reperfusion injury and improves transplant outcomes. Preclinical studies have shown that DHOPE can safely extend the preservation of donor livers for up to 24 h. Methods: We conducted an IDEAL stage 2 prospective clinical trial comparing prolonged (≥4 h) DHOPE to conventional (1–2 h) DHOPE for brain-dead donor livers, enabling transplantation the following morning. Liver allocation to each group was based on donor hepatectomy end times. The primary safety endpoint was a composite of all serious adverse events (SAE) within 30 days after transplantation. The primary feasibility endpoint was defined as the number of patients assigned and successfully receiving a prolonged DHOPE-perfused liver graft. Trial registration at: WHO International Clinical Trial Registry Platform, number NL8740. Findings: Between November 1, 2020 and July 16, 2022, 24 patients were enrolled. The median preservation time was 14.5 h (interquartile range [IQR], 13.9–15.5) for the prolonged group (n = 12) and 7.9 h (IQR, 7.6–8.6) for the control group (n = 12; p = 0.01). In each group, three patients (25%; 95% CI 3.9–46%, p = 1) experienced a SAE. Markers of ischemia-reperfusion injury and oxidative stress in both perfusate and recipients were consistently low and showed no notable discrepancies between the two groups. All patients assigned to either the prolonged group or control group successfully received a liver graft perfused with either prolonged DHOPE or control DHOPE, respectively. Interpretation: This first-in-human clinical trial demonstrates the safety and feasibility of DHOPE in prolonging the preservation time of donor livers to enable daytime transplantation. The ability to extend the preservation window to up to 20 h using hypothermic oxygenated machine preservation at a 10 °C temperature has the potential to reshape the landscape of liver transplantation. Funding: University Medical Center Groningen, the Netherlands.</p

Machine Perfusion of Donation After Circulatory Death Liver and Lungs Before Combined Liver-lung Transplantation

Shortage of deceased donor organs for transplantation has led to the increased use of organs from donation after circulatory death (DCD) donors. There are currently no reports describing outcomes after multiorgan transplantation with DCD livers. The use of DCD organs for multiorgan transplantation can be enhanced if the detrimental effects of prolonged cold ischemia and subsequent ischemia-reperfusion injury are overcome. We present a case in which the liver and lungs of a DCD donor were preserved using ex situ machine perfusion for combined liver-lung transplantation. The recipient was a 19-year-old male patient requiring bilateral lung transplantation for severe progressive pleural parenchymal fibroelastosis and portal hypertension with portal vein thrombosis. The donor liver was preserved with dual hypothermic oxygenated machine perfusion, whereas the lungs were perfused using ex vivo lung perfusion. With ex vivo lung perfusion, total preservation time of right and left lung reached 17 and 21 h, respectively. Now, 2 y after transplantation, liver function is normal and lung function is improving. To conclude, we suggest that combined transplantation of DCD liver and lungs is feasible when cold ischemia is reduced with ex situ machine perfusion preservation