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The regulatory acceptance of xenotransplantation is an essential step in realizing the most promising technology for restoring normoglycemia to people with diabetes. There have recently been some very positive developments that have demonstrated the safety of xenotransplantation. Some of these articles discuss the risks associated with using tissue from transgenic pigs and the use of immunosuppression. In the case of islet xenotransplantation, neither transgenic pigs nor immunosuppression is used, making this treatment safer than many everyday medical procedures.
 
New Scientist -- CDC Report on Recipients of Porcine Tissue Transplants (August 8 1998)
New Scientist -- Editorial on Xenotransplantation (August 8 1998)
BBC News Online -- Pig Viruses Don't Pass to Humans (August 8 1998)
XVII World Congress of the Transplantation Society, Montréal (July 12-17 1998)
Bioartificial Organs II: Conference Report, Banff, Alberta, Canada (July 18–22 1998)
 
News
8 August 1998 Vol. 159 No. 2146
So far, so good...
 By Andy Coghlan
fears that novel viruses might wreak havoc in transplant patients who receive pig organs may be groundless, according to findings due to be presented this week in London. The findings, based on screening samples from patients UK guidelines for xenotransplantsexposed to pig tissue, provide the first compelling evidence that dormant pig viruses do not spread to humans, causing new and incurable infectious diseases. 

 This doomsday scenario has been the main obstacle holding up clinical trials to test "xenotransplants" of pig organs. Robin Weiss of the Institute of Cancer Research in London raised the fears last year when he showed that two pig viruses could spread from pig to human cells in the lab. 

 The viruses, called porcine endogenous retroviruses, have become part of the pigs' own DNA over millennia. Though dormant in the pigs, Weiss's experiments highlighted the possibility that the viruses might reawaken in humans. 

 In the light of Weiss's findings, the US Food and Drug Administration (FDA) temporarily withdrew approval for clinical trials related to xenotransplantation. But since analysing the outcomes of earlier trials, several groups of researchers in the US are starting to dispel the fears. 

 Some of their results were due to be announced this week in London at a closed workshop on pig viruses organised by the UK Xenotransplantation Interim Regulatory Authority, which oversees xenotransplantation proposals in Britain. Last week, it launched guidelines for xenotransplant experiments (see Table). 

 At the meeting, Walid Heinene and Louisa Chapman of the US Centers for Disease Control in Atlanta, Georgia, were due to describe their study of 10 Swedish diabetic patients who received transplants of pig pancreatic islet cells, which manufacture insulin. Even in patients where the cells survived for more than a year, the researchers found no pig virus DNA. Nor could they find any antibodies against pig viruses in their blood, suggesting the blood was free of infection. And there were no signs of viral reverse transcriptase enzymes, a telltale sign of any unidentified viruses. 

 "We looked in blood, lymphocytes and serum, and found nothing," says Heinene. "These are reassuring data." But he cautions against misplaced optimism: "How much we can extrapolate to other types of transplant is not known." 

 Diacrin, a company in Charlestown, Massachusetts, was also due to present encouraging results this week. It has developed treatments for Parkinson's disease and Huntington's disease using brain cells from pig fetuses. The company did not find pig viruses in any of its 24 patients, including one treated three years ago.  

Other companies reported similar results last month in Montreal at the World Congress of the Transplantation Society, an international organisation. They included Circe Biomedical in Lexington, Massachusetts, a company that has used pig liver cells in a device that extracts toxins from the blood of patients with liver failure. Of the 25 traceable patients receiving the treatment, none has acquired pig viruses. Last week, following approval from the FDA, Circe resumed trials in the US and Europe. 

 Although the negative results for transmission of the viruses are encouraging, caution remains. "Any data that are negative are great, and they mean we can move to the next level of evaluation," says Jon Allen of the Southwest Foundation for Biomedical Research in San Antonio, Texas. But what happens in people who receive an entire organ might be very different, he says. 

 Weiss agrees. "If there are negative results does it mean no one's infected, or that you can't detect it?" he asks. "It's fraught with difficulties." 

 related sites: 
  

  • Xenotransplantation: what, why, which species...  
  • First East Coast Patient Treated With Bioartificial Liver  
  • Northwestern Joins Baxter In Transgenic Pig Liver Trial  
  • Working groups develop policies on xenotransplantation--Focus  
  • Transplantation Society--XVII World Congress
  •   
    From New Scientist, 8 August 1998 
    © Copyright New Scientist, RBI Limited 1998

     

  • Xenotransplantation - Spectrum of Safety and Complexity (Source: TIF)
  • EDITORIAL
    EDITORIAL GRAPHIC
     A double-edged sword
    who on earth would choose to have a pig's heart transplanted into them if they could avoid it? The jokes would be bad enough. Just think: every time you ate too much or answered a question with a grunt, some clever devil would be ready with a merry quip. And the current fashion for wearing a ring though one nostril would definitely die a death.  

    But like it or not, we are probably going to need hearts and other organs from animals to replace and repair human ones. The wait for humans organs for transplants is already so long that many patients die before a suitable donor turns up. Animal organs could fill the gap before, in the more distant future, we come up with cleverer ways of healing damaged organs.  

    So far, research aimed at making xenotransplantation possible has had some wild ups and downs. All attempts to transplant whole animal organs, from the first operations in France in the 1900s to those in the US in the 1990s have failed. Optimism returned in 1994 when one biotech company showed that it was possible to genetically engineer pigs so that their organs could avoid immediate immune ejection if transplanted into a human. But the prospects soon worsened again as the risks of animal to human transplants became more apparent.  

    It is easy to understand why. Pigs and other animals carry viruses hidden within them. How do we know that one of them might not turn out to be like HIV or other viruses that trigger devastating epidemics when they pass from animals to humans? Sceptics point out that man has lived for so long with pigs that we should have encountered all the pig viruses by now. And others point out that surgeons have been using valves from pigs' hearts to replace human ones for 25 years.  

    Unfortunately, neither argument is convincing. Viruses have unique opportunities to infect humans if they are in an organ inside the human body--especially if the individual is taking immunosuppressive drugs. Valves from pigs' hearts are irrelevant to the argument. They are soaked in glutaraldehyde before use and are no more alive than a toenail.  

    The reality of the risks was driven home last year. Robin Weiss of the Institute of Cancer Research in London and David Onions at the University of Glasgow showed that pig retroviruses could indeed infect human cells in a laboratory culture. Pessimism reigned, and the US Food and Drug Administration promptly put a stop to any pig organ transplants until more was known.  

    This week, we're back to optimism with a new study which shows that people who have received pig tissue as skin grafts and to repair parts of the pancreas and liver do not pick up any pig viruses (see this week's news section). But before we get too excited, let's remember how far we still are from a real assessment of the risks of xenotransplantation.  

    The new studies come from people who had received small amounts of living tissue. A whole organ provides a much bigger reservoir of viruses. And there are special risks because the organ would have come from a pig that had been genetically altered to make it more compatible with a human.  

    This process could play into the hands of viruses. There are two main ways that a pig can be genetically altered to improve its compatibility. One aims at changing chemicals on the surface of pig cells so that when they are grafted into a human they look more like human cells and do not trigger the hyperacute immune reaction that quickly destroys them. The other aims at adding genes that help to block the hyperacute reaction.  

    When viruses form, they wrap themselves up with a coat built within the cell in which they live. There is a chance that viruses lurking in genetically engineered pigs could wrap themselves in a coat that also made them look more human and thus avoid the immune system. Or they might acquire the power to weaken the reaction against them.  

    It would be a tragedy if genetically engineered pigs designed to have organs that fool the human immune system also produced viruses that fool the human immune system. It might not happen but despite the upbeat news this week xenotransplantation of an entire organ needs to be handled with extreme caution. Nothing could be worse than if in trying to save our lives we inadvertently engineered a plague.  

    From New Scientist, 8 August 1998 

    © Copyright New Scientist, RBI Limited 1998
    Pig Viruses 'Don't Pass to Humans'
    BBC News Online (08/08/98)

         A New Scientist review of two U.S. studies has concluded that the possibility of pig viruses being transmitted to humans via xenotransplants "may be groundless."  The first study, conducted by the Centers for Disease Control and Prevention and to be announced this week at a meeting of the U.K. Xenotransplantation Interim Regulatory Authority, found that 10 Swedish diabetics who were given pig pancreatic cells have not subsequently become infected with tainted pig DNA.  The second investigation, conducted by the U.S. firm Diacrin, reportedly found that none of 24 patients with Parkinson's and Huntingdon's disease who received fetal pig cells had become infected with pig viruses. Despite its findings, the New Scientist conceded that
    transplanting whole organs could hold a much higher risk than transplants of cells.

    Trip Report
    XVII World Congress of the Transplantation Society
    Montréal, July 12-17 1998
    André La Prairie, Policy Division, TPP
    Heath Canada

    The Transplantation Society was founded in 1966 to act as "the principle forum for the advancement of basic and clinical transplantation throughout the world". This year more than 3,700 attendees from over 40 different countries participated in the Transplantation Society’s 17th World Congress in Montréal, which boasted over 1700 symposia, oral and poster abstracts addressing areas such as molecular biology, gene therapy, tissue engineering, xenotransplantation, ethics and economics.

    The emphasis of this congress was science. Plenary sessions were predominantly on topics such as T-cell activation, molecular modelling, and cytokine signalling pathways. Breakout sessions also were heavy on immunobiology, chronic rejection and immunotolerance - and most presentations/posters compared and contrasted various regimes of immune suppression therapies.

    But the non-molecular scientist could not complain. Without the science and plethora of drug choices for immune suppression there would be no transplantation and no meeting. The keynote session emphasised this reality. Titled a "Half Century Retrospective of Transplantation as Viewed by the Protagonists" it was the ultimate panel discussion on transplantation. Every major immunologist or transplant pioneer was present: Fritz Bach, Leslie Brent, Sir Roy Calne, Jean Dausset, James Hardy, Lady Jean Medawar, Joseph Murray, Felix Milgrom, John Najarian, Felix Rapaport, Robert Schwartz, Norman Shumway, Thomas Starzl, and Paul Terasaki were invited to present their perspective of the first 50 years of transplantation. Someone was both brilliant and cruel in coming up with a 4 minute maximum presentation time for each panel member with an automatic microphone shut off. Each member could easily have kept the audience captivated with stories of how the many secrets of rejection were deciphered and why transplantation has become a routine clinical reality today. In fact it is the success of these pioneers and many others that has resulted in the many outstanding issues transplantation now faces: exponential increases in demand for organs, new transplant therapies for disease treatment, use of marginal donors and organs, living non-related donors, controversial listing and allocation practices, sale of organs and .... xenotransplantation. Yesterday’s protagonists may not be able to help with these new problems. In fact when asked for a vision of what transplantation would be like in another fifty years, most declined the opportunity to speak.

    The media attention and daily press releases from the congress also emphasised the hightened public interest in transplanation. A week before the congress the Lancet published an article by a group of well known scientists suggesting that the sale of organs may be acceptable. The Canadian Organ Replacement Registry released its annual report, confirming that organ donation in Canada continues to decrease and waiting lists continue to rise. The plenary session on Friday focused on the possibilities of xenotransplanation and every day the media released stories focusing on the lack of available organs for transplant.

    To review the many press releases and reports from the congress, visit the 1998 congress webpage.

    Some issues and observations:

    Expanding donor options: the use of marginal donors and marginal donor organs. Many different options were explored in a series of symposia on the ways to increase the number of available organs for transplantation. Expanding acceptable donor criteria to include the older and younger donors, use of hepC positive donors (into HepC positive recipients), cleavage of organs (liver/lung/pancreas) to double the number of organs, use of living non-related donors, organ size reduction, diabetic and hypertensive donors and use of the non-heart beating donor were discussed. All these options can increase the number of available donors. But suboptimal donors mean suboptimal organs. And suboptimal organs suggest lower graft survival and increased risk to the recipient. Clearly guidelines will have to be established for the acceptable use of these organs. Informed consent for suboptimal organs was raised, but not well recieved by the majority of clinically focused transplanters.

    New paradigms in graft loss: While recent efforts to improve long term graft survival have focused on powerful new immuno-pharmacology options to the patient at time of transplant, new evidence suggests that the pathophysiology of donor organs at time of brain death may also be significant. This suggests that important information about the donor and organ needs to be collected (ischemic time, cause of death, hyper/hypotension, reperfusion injury...) and made part of national registries - esp with the trend towards expanding the donor pool to include marginal donors, non-heart beating donors and longer cold storage times.

    Ethics, ethics, ethics.... There were some wonderful presentations in the breakout sessions and the poster presentations on ethics. Organ rationing, informed consent, use of marginal donors, economics and commerce, and presumed consent were all touched on. But you can’t help think that more emphasis and a plenary session could have been given to this area. Clearly the media interest and the congress spokespersons were eager to discuss donor shortages, the issue of exchanging money for organs, and stragegies to improve awareness and professional support.

    Xenotransplantation: A number of breakout sessions and a major plenary session addressed xenotransplantation specifically, as well as presentations under cell transplantation and tissue engineering. Louisa Chapman’s (CDC) presentation on xenozoonoses was the best of a very interesting cache of presentors. (Ian Kennedy’s presentation on Guidelines for Regulation was a close second, but he was too quick to blame all of Canada for his powerpoint slide presentation problems). Most of the focus on xeno at this congress was on the biology of rejection and potential hazards from pathogen transmission. A significant paper by Heneine et al described methods of PCR surveillance on 10 diabetic recipients of pig islet cells. No evidence of PERV transmission was observed, but the authors noted this was a small trial and results are specific for this patient population and the specific transplant regime.

    While some authors touched on the future issues of xenotransplantation (inferior to allotransplants, lifelong monitoring, monitoring of close contacts....) very little was said about the social and public issues of xenotransplantation. Even Fritz Bach avoided the use of the word "moratorium" when he discussed the possiblilities of xenotransplantation in his acceptance speech as a Medawar award recipient. (Drs . Anthony P. Monaco and Felix T. Rapaport - both transplant surgery professors - were also recipients of the Medawar award).

    In recognizing the importance of this emerging field, the Transplantation Society established a sub-committee on xenotransplantation. And Japan will be hosting the 5th international congress on xenotransplantation in 1999. (For more info, go to http://www.pin-japan.com/xeno)

    Other interesting items: The Canadian Transplantation Society is changing its name to the Canadian Society for Transplantation (CST). This is because of the confusion with the Canadian Thoracic Society, which is also known as CTS. The CST and the Canadian Association of Transplantation (coordinators, allied health professionals) will continue with their joint annual meetings next year, but will not link with the Royal College meeting. Instead they will partner with the Canadian Immunology Society. Joint symposia and the popular transplant debates will continue to be the highlight of this joint meeting.

    The Kidney Foundation of Canada’s "Links to Success" advisory committee has proposed a special workshop to determine a framework for donation and awareness in Canada, with required referal as one of the possible focus elements. The KFC will consider this proposal and look for funding options. The framework could be forwarded to the Canadian Advisory Committee on Health Service’s federal/provincial steering committee addressing organ/tissue donation and distribution. The KFC advisory committee strongly endorsed the need to include tissue donation in any donation strategy.

    The Roche Organ Transplantation Research Foundation was announced at the congress, setting aside 25 million Swiss Francs for transplantation research.

    The conference organizing committee (Guttman, Daloze et al) did a great job of putting together the largest congress ever of the Transplantation Society. Montreal was the perfect Canadian city to hold this meeting. Not only were the restaurants and social events a good backdrop to the meeting, but 40 years ago (1958) Montreal’s Royal Victoria Hospital was the site of Canada’s first kidney transplant. The Palais des Congres was an excellent facility to hold the large plenary and multiple breakout sessions, and the timing and access to all sessions was very smooth. NEXT CONGRESS: Rome, 2000.

    [This report represents the opinions of the author - A. La Prairie - and not those of the Therapeutic Products Programme or Health Canada.]

     

    Bioartificial Organs II: Conference Report
    Cosponsored
    The Engineering Foundation
    The Juvenile Diabetes Foundation International
    Banff, Alberta, Canada, July 18–22, 1998
    July 28, 1998
    David Hunkeler
    Artur Bartkowiak
    The Bioartificial Organs II Conference, which addressed the problems at the interface of encapsulation technology, novel materials and medicine, was attended by 102 participants from 15 countries. There were a total of 10 sessions, including one dedicated to posters, with 60 papers presented. This report highlights the innovative advances that have occurred over the past two years. Copies of the abstracts are available from the Engineering Foundation.

    Opening Lectures
    Joseph Kennedy (U. Akron) gave the first lecture and discussed novel biomaterials based on polyisobutylene, which is sterilizable and FDA approved. By varying the solvent the characteristics of the network would be changed from hydrophilic to hydrophobic, with one microphase dispersed in the other, or a true amphiphilic structure. He referred to this as a “chameleon.” The main application, which has been patented, is the production of hollow fibers for islet cell isolation. Kennedy has won both the pure and applied polymer science awards from the ACS, their highest macromolecular honors. He will visit the EPFL in 1999. Fritz Bach (Harvard) discussed the ethics of xenotransplantation. He had four main points:
     

    1. The risk to the public should determine the public mechanism to deal with and regulate a technology.
    2. Individual informed consent must be modified to include patients’ close contacts.
    3. The significant public risk implies that an interactive regulation will be required.
    4. Viruses can return to the animal donors after an incubation period in humans.
    Bach does not believe in the Swiss democratic model and has proposed a national “stakeholder” committee to “represent” the public. However, his list of representatives would be much more inclusive of experts rather than the family unit.

    Plenary Session 1: Clinical Applications and Novel Materials
    Thomas Chang, from McGill, discussed Artificial Cells, Immobilization and Encapsulation. He noted that 85% of the 400,000 people per annum suffering from chronic renal failure die since they cannot afford dialysis treatments. Therefore, he has constructed an artificial cell which contains an urease enzyme which converts urea to ammonium. He also briefly discussed a modified hemoglobin blood substitute which is in phase II/III clinical trails. The latter is discussed in the July 1998 issue of Nature Biotechnology.

    Patricia Chang (McMaster) gave a nice lecture on the use of encapsulation in gene therapy with the application to treat lyposomal storage diseases. Artur had long discussions with Professor Chang, who is interested in a cooperation.

    Ray Rajotte discussed his unique method for islet cryopreservation. He prefers a slow freeze-thaw protocol with a slow cool (0.25 °C/min) to °40 or –75 °C, then a rapid plunge to –116 °C. DMSO is added as a cryoprotectant in a stepwise manner. This is carried out at 20 °C to permit time for the DMSO to penetrate. A “nucleation” step is also needed. This involves the supercooling of test tubes to –7.5 °C followed by nucleation with a small piece of ice until of the latent heat of fusion is given off. The rapid thaw involves a heating of 200 °C/min to minimize the time for ice crystals to grow and lyse the cells. Their yield is approximately 80%, three times higher than anyone else. The recovery increases to 88% if encapsulated in an Alginate-Poly-L-Lysine capsule. They have transplanted islets into the kidney and have some animals 5 to 7 years post transplant. He is presently in the clinic.

    Yoshihito Osada (Sapporo) presented a talk similar to the one given at MacroHavana’97. His latest advance is an ability to convert chemical energy into electric power. He also, in general, is working on artificial muscles and the artificial bladder. His advances have come by observing nature. For example, he has hydrogels which can swim, and this is based on observations that the drag coefficient in motion of fish is reduced with the size, or speed (i.e., Reynolds number) of the animal.

    Session 2: Bioartificial Pancreas
    Anthony Sun (U. Toronto) mentioned that he uses 300-micrometer capsules, which were optimized by Maleki in 1987. Islets are isolated using Calafiore’s method, and he noted that they can obtain good quality islet tissue from 25% of the pigs harvested. In his large animal monkey trials 7 of the past 10 have been insulin free, one for seven years. The transplanted tissue corresponded to 10–15,000 islets per kg of weight. The monkeys receive a second dose of capsule when their glucose increases. The have observed insulin in glucose tolerance tests, with triglycerides and cholesterol decreasing to normal levels three months after transplantation. At present they are working on clinical studies in China. Sun mentioned that the source of spontaneously diabetic mice is very limited and the fact that other groups, which he cannot identify, are obtaining the majority of the supply has hindered his research. The potential exclusion from a key market is a reason we must always consider. The most significant immediate implication is the inability to use Pronova alginate to a threat for an ultimate production stoppage.

    Riccardo Calafiori (Perugia) discussed the advantages of alginate-poly-L-orthine membranes, including a tighter MWCO. He believes the future is with the “medium-sized capsules” of 350 µm. This would correspond to a transplant volume of 35 mL for a 70 kg individual. He has observed five-month diabetes reversal in dogs and prefers transplantation to the peritoneum or omental pouch. He noted that pig islet diameter and quality was a very strong function of the type of pig herd, with Yorkshire Hampshire preferred. While pig islets dissolve in culture in 4 days, they remain for weeks if immobilized.

    Michael Dornish of Pronova gave an lecture on the proposed “standardization” of biomaterials.

    Phillipe Morel (HUG) discussed clinical islet transplantation. He noted that Roche’s new “Liberase” enzyme does not work as well as the former collagenase. However, Rajotte warned that the liberase required islet processing modifications and could work. The results of clinical transplantation studies are that 25% of individuals are glucose intolerant after 5 months with 50% insulin independent after 5 years. Morel concluded that pancreas transplantation works, but there is a high price to pay (morbidity of 100% and mortality of 10%). Islet transplantation results show C-peptide levels of >0.5 after one month in 70% of patients with 10% of patients off insulin for one week (these are metrics). Usually, the transplants are in combination with a kidney or, preferably, after a kidney. Ten institutes in the world carry out clinical islet transplantation. He concluded that long-term insulin independence can be achieved in autografts, though with allografts the results have not been successful. He recommends the improvement in islet isolation, the development of better enzymes (liberase does not work!) and the evaluation of the effect of endotoxin contamination on islet quality. He noted that a new idea in islet isolation could be to destroy the exocrine and collect the endocrine. Bach said this was the most impressive data he had seen.

    Session 3: Tissue Sources, Xeno- versus Allografts
    Simon C. Robson (Beth Israel Deaconess Medical Center, Boston) discussed the factors in xenograft rejection. He categorized rejection factors into the two groups, hyperacute rejection (HAR) and delayed xenograft rejection (DXR). Analysis of the factors underlying DXR should lead to the development of genetic approaches that, in conjunction with selected pharmacological means, may further prolong xenograft survival to a clinically relevant extent.

    Colin Weber (Emory University, Atlanta) discussed the techniques for prolongation of islet xenograft survival. NOD immunoreactivity toward encapsulated xenogenic islets was found to be helper-T-cell-dependent (this dates to work carried out in the late 1980s). Xenogenic islet microencapsulation plus host immunomodulation (NOD mouse treatment with CTLA4lg, which interferes with co-stimulatory interactions between antigen presenting cells and T-cells) promotes long-term islet xenograft graft survival.

    Gregory S. Korbutt (University of Alberta, Canada) presented the paper on neonatal porcine as a source of tissue for islets. The neonatal porcine pancreas can be used for the isolation of a large number of viable islet cells, and due to their unlimited supply and inherent capacity to proliferate and differentiate, they constitute an attractive source of insulin-producing tissue for clinical transplantation. He will participate in BIO+AO III.

    Bernard Thorens discussed the encapsulation of genetically engineered cells secreting glucagon-like peptides for the treatment of non–insulin-dependent diabetes (Type II). They encapsulate in a polyethersulfone membrane with a MWCO of 280,000 (Clark Colton later noted that MWCO was an ill-defined term that should not be used, and there was a large consensus in agreement with him). They implanted the GLP-1 cells subcutaneously and reported positive results in high-fat fed mice. The next step is to go into the intraperitoneum.

    Session 4: Processing and Technology
    Michael Sefton (U. Toronto) discussed a novel method for conformal HEMA-MMA capsules where the islet was centrifuged through a gradient, one layer of which was the polymer. He noted that you could centrifuge either upwards or down the density gradients. He concluded that in vitro response was evident and that organic solvents were possible with islets. One problem encountered is that the PEG200 used to dissolve the HEMA-MMA copolymer dehydrates the cells, imparting a type of “toxicity.”

    Clark Colton discussed the macrodevice developed by Baxter and being tested by C. Gordon Weir. It is a planar device that uses alginate to immobilize islets. Diabetes reversal has been reported for a few weeks. Oxygen is a problem, and he noted that tissue survivability increased if the capillary spacing was 100 µm or less. Clearly they prefer vascularization, and they are using VegF to induce blood vessel formation. They noted that they intend to generate oxygen locally to prevent necrosis, for example, using electrochemical catalytic means.

    Rebecca Li from Cytotherapeutics discussed matrices for cell attachment and noted that agarose, alginate, chitosan, PEO, polyvinyl alcohol and polyethylene had been reported in the literature. They employ PAN/PVC tubes with a diameter of 100 µm. They observe necrosis in six weeks with PC-12 cells in chitosan. They are in phase II clinical trials in the US, Switzerland and Central Europe.

    Lunch Talk: “Portal or Peripheral Insulin Delivery: Which Is Better?”
    Allan Cherrington gave an excellent lecture on the role of two hormones, insulin and glucagon, on the carbohydrate metabolism. His studies, which were done on living dogs, examined the sensitivity of the alpha and beta cells of the pancreas to changes in the concentration of plasma glucose. In vivo the beta cell is extraordinarily sensitive to glucose, which is why the blood sugar level is so highly controlled in non-diabetics. To summarize a long talk, Allan reported that the liver responds quickly and sensitively to insulin with complete inhibition of glucagon breakdown. He recommends delivery of insulin portally (portal vein of liver), which has obvious implications on the site of islet, or encapsulated islet, transplantation. Glucagon has the opposite effect of insulin and, to some extent, these two hormones battle. It is also very sensitive. Peripheral insulin, by comparison, results in large swings in blood sugar levels. It is clear that a bioartificial pancreas will be, in terms of performance, between the portal and peripheral insulin delivery.

    Session 5a: Panel Discussion on Scale-Up and Related Issues
    David Scharp of Neocrin reported on the scale-up of Hubbell’s conformal PEG coating process. They can now encapsulate 20,000 islets per minute and Scharp, a surgeon, does not see technology as a limitation to the bioartificial pancreas. The major scale-up item for him is the tissue source, with quality control, cost effectiveness and the production of uniform MWCOs as other issues. They report positive results (porcine to rat xenograft) for 200 days, with C-peptide measured for >3 months. Some sobering statistics include that fact te 40,000 pigs will be needed if 4,000 encapsulated islet transplantations are to be performed per year (Robert Lanza from BioHybrid Technologies questioned how these carcasses would be disposed of). This would result in a tissue cost of $3000/transplant. Neocrin has three herds separated by several kilometers: a stock herd, breeding herd and nursery/finishing herd. Animals are transported from the midwest to California where they are quarantined. At present, pancreas digestion remains a batch process.

    Dawn Applegate from Advanced Tissue Sciences mentioned that the limit a tissue can be separated from an oxygen source is 250 µm. Problems in reactor design include the pulsatile flow, the need for aseptic conditions for up to two months and the cryopreservation of large amounts of tissue simultaneously. She noted that it took ATS eight years to put their bioartificial skin on the market. Companies, therefore, have a strategy to start clinical trials prior to proof of concept.

    The discussion concluded with a consensus that metrics need not be fundamental but only representative of in vivo effects.

    Session 5b: Oral and Craniofacial Related Issues
    Four excellent lectures will be summarized by Eleni Kouselvari (NIH).

    Poster Session
    During the session 20 posters were presented, which can be divided into the 4 groups:
     

    1. Materials for bioartificial organs (preparation and sterilization: 3 posters)
    2. Microcapsules and microspheres as bioartificial organs. A total of 7 posters described the following systems:
    3. Bioreactors as bioartificial liver (3 posters)
    4. Tissue for bioartificial organs. This included methods of isolation, purification and characterization of activity (7 posters)
    News:
    J. P. Kennedy presented a new fully synthetic biocompatible and biostable rubbery network. These amphiphilic networks were created by copolymerization of methacrylate-telechelic polyisobutylene with various water soluble methacrylates. This material can be used to prepare small cylindrical tubes and used for macroencapsulation of living cells.

    Session 6: Molecular Aspects in the Prevention of Recognition and New Biomaterials
    Lola Reid (University of North Carolina) gave a nice lecture on cell matrix and gene expression. She mentioned that heparin and heparin-sulfate are good candidates as materials for bioartificial pancreas due to their high biocompatibility and insulin non-interaction character. She also noted that insulin of non-host structure (xenotransplantation) can create host immunoresponse and in the consequence the transplant rejection.

    Ron Gill (Univ. Colorado) gave an excellent talk on antigen pathways in allograft and xenograft rejection. His main conclusion, which is important for our work, is that for an allograft it is the graft that is rejected, while for a xenograft it is the host that is the most important.

    Tom Cavanaugh (Roche/Boehringer Mannheim) discussed enzyme sources for tissue “de-engineering” and dissociation. He noted that collagenase is purified by affinity chromatography, as well as anion and cation exchange. He claimed that liberase provided better batch-to-batch reproducibility and low endotoxin levels (<50 EU/mL). They work with Rajotte, Calafiori as well as groups in Arizona and Minneapolis. Although not mentioned they must supply Riccordi in Miami. He reported improved results in C-peptide levels after one month in more recent human allografts (unencapsulated). However, the conclusion was that tissue dissociation can cause cell damage.

    Hiroo Iwata discussed a multicomponent capsule very much like the Prokop/Hunkeler/Wang material, which includes an agarose/polystyrene sulfonate blend coated with the oligomeric polybrene to form a complexed membrane with CMC on the outside. The agarose substitutes for our alginate and serves to increase viscosity, while the PSS is reactive. He claimed that this capsule regulates compliment function in hamster-to-mouse xenografts.

    Paul Dubin (Purdue) discussed coacervation from a physical chemical perspective. He noted that it is the local charge that is important in complexation, implying the complimentarity of charge spacing between oppositely charged polyelectrolytes is a key parameter. He also noted that the micro and macro viscosities can differ by two orders of magnitude.

    Session 7: Bioartificial Liver, Implants and Extracorpeal Cellular Devices
    Claudy Mullon (Circe Biomedical) discussed extracorpeal liver support devices. Their HepatAssist device is used as a bridge to a whole organ transplant and the results are encouraging with over 90% of patients having partial hepatic failure surviving for 30 days.

    Jorg Gerlach discussed the development of new bioreactor construction containing pig liver cells to act as hybrid human liver support system. The primary human, as well as porcine, hepatic cells spontaneously reestablish to aggregates which immobilize themselves between the membranes of bioreactor. Such bioreactor-containing porcine liver cells were used clinically as a bridge to liver transplantation in six patients with acute liver failure.

    Jack Patzer (Pittsburgh) discussed a competing device to the HepatAssist. They use a membrane with a MWCO of 100,000 daltons. The preliminary work will go to Phase I clinical trials.

    Achilles Demetriou (UCLA) discussed clinical experience with the extracorpeal BAL. With positive results they will move to Phase II/III clinical trails. It is important to note that the BAL is a macro device contained outside the body. It involves perifusion of a patient’s blood with cells and does not involve immunoisolation.

    Session 8: Bioartificial Skin, Bone and Cartilage
    Jiri Prenosil (ETHZ) discussed a novel controlled bioreactor for skin grafts. The “KERATOR” is automatic and can produce 0.5 m² of skin in 2 weeks. This would lower hospital stays and costs from $300,000 to $100,000 for a typical patient. The reactor is based on a cultured keratinocyte autograft.

    Kermit Borland (Reprogenesis) discussed injectable hydrogels containing autologous chondrocytes as engineered tissues. They are currently in Phase III clinical trials for bioartificial cartilage. Their procedure involves the removal of a 5x5-mm piece of cartilage from the back of the ear, culturing, mixing with alginate and injection into the patient (20 million cells/cm³). They use Pronova alginate and gel it with calcium. The viscosity-time curve follows an S-shaped curve. Rapid cell infiltration is observed. The main applications include vesicoureteral reflux (reversal of urine flow from bladder to ureter) and incontinence (loss of mechanical integrity of tissue of ureter).

    Hassan Uludag (Genetics Institute, now at U. Alberta) discussed bone induction. Specifically, collagen sponges with bone morphogenic proteins (BMPs) were evaluated pharmacokinetically. The isoelectric point is the main factor controlling release. Sterilization was via EO of the whole collagen sponge. Gamma radiation was not evaluated. The talk was excellent.

    Gail Naughton (President, Advanced Tissue Sciences) concluded the conference, as she did BIO+AO I, with a talk on bioartificial cartilage. They have designed bioreactors to provide uniform cell growth on a PGA scaffold. They are able to preserve the cartilage at 4 °C for 12 weeks. Sheep will be their next model. The future is an artificial joint, which includes both cartilage and bone grafts. A question was posed, after the session, if an artificial uterus would someday be in the works.

    Contact
    David Hunkeler
    Laboratory of Polymers and Biomaterials
    Swiss Federal Institute of Technology, CH-1015 Lausanne, Switzerland
    T: 41-21-693-3114; F: 41-21-693-3680;
    E-mail: hunkeler@igc.dc.epfl.ch
     

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