Lars Burdorf

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.).

We describe the incidence of early graft failure (EGF, defined as loss of function from any cause within 3 days after transplant) in a large cohort of GalTKO pig organs transplanted into baboons in three centers, and the effect of additional expression of a human complement pathway-regulatory protein, CD46 or CD55 (GalTKO.hCPRP). Baboon recipients of life-supporting GalTKO kidney (n = 7) or heterotopic heart (n = 14) grafts received either no immunosuppression (n = 4), or one of several partial or full immunosuppressive regimens (n = 17). Fourteen additional baboons received a GalTKO.hCPRP kidney (n = 5) or heart (n = 9) and similar treatment regimens. Immunologic, pathologic, and coagulation parameters were measured at frequent intervals. EGF of GalTKO organs occurred in 9/21 baboons (43%). hCPRP expression reduced the GalTKO EGF incidence to 7% (1/14; P < 0.01 vs. GalTKO alone). At 30 mins, complement deposits were more intense in organs in which EGF developed (P < 0.005). The intensity of peri-transplant platelet activation (as β-thromboglobulin release) correlated with EGF, as did the cumulative coagulation score (P < 0.01). We conclude that (i) the transgenic expression of a hCPRP on the vascular endothelium of a GalTKO pig reduces the incidence of EGF and reduces complement deposition, (ii) complement deposition and platelet activation correlate with early GalTKO organ failure, and (iii) the expression of a hCPRP reduces EGF but does not prevent systemic coagulation activation. Additional strategies will be required to control coagulation activation.

BACKGROUND: The purpose of the present study was to examine whether the effect of coronary stenoses of variable severity on myocardial perfusion can be quantitatively assessed in vivo by analysis of fluorescent cardiac imaging (FCI) compared with the gold standard, the fluorescent microsphere method. FCI is a novel technology to visualize coronary vessels and myocardial perfusion intraoperatively using the indocyanine green dye with an infrared-sensitive imaging device. METHODS AND RESULTS: Graded stenoses and total vessel occlusion of the left anterior descending coronary artery were created in 11 open-chest pigs. Stenoses were graded to reduce resting left anterior descending coronary artery flow by 25%, 50%, 75%, and 100% of baseline flow measured by transit-time flowmeter. FCI images were analyzed with a digital image processing system. The impairment of myocardial perfusion was quantified by background-subtracted peak fluorescence intensity and slope of fluorescence intensity obtained with FCI and compared with myocardial blood flow assessed by fluorescent microsphere. All stenoses resulted in an impairment of myocardial perfusion visualized by FCI. Occlusion of the left anterior descending coronary artery resulted in a total perfusion defect (no fluorescence intensity) of the corresponding anterior myocardial wall. During graded stenosis and total vessel occlusion, normalized background-subtracted peak fluorescence intensity and slope of fluorescence intensity decreased significantly (P<0.0001). Both background-subtracted peak fluorescence intensity (r=0.92, P<0.0001) and slope of fluorescence intensity (r=0.93, P<0.0001) analyzed by FCI demonstrated good linear correlation with fluorescent microsphere-derived myocardial blood flow. CONCLUSIONS: The impairment of myocardial perfusion in response to increased coronary stenosis severity and total vessel occlusion can be quantitatively assessed by FCI and correlates well with results obtained by fluorescent microsphere.

Consistent survival of life-supporting pig heart xenograft recipients beyond 90 days was recently reported using genetically modified pigs and a clinically applicable drug treatment regimen. If this remarkable achievement proves reproducible, published benchmarks for clinical translation of cardiac xenografts appear to be within reach. Key mechanistic insights are summarized here that informed recent pig design and therapeutic choices, which together appear likely to enable early clinical translation.