A New Enzyme Mixture to Increase the Yield and Transplant Rate of Autologous and Allogeneic Human Islet Products

Abstract

Background The optimal enzyme blend which maximizes human islets yield for transplantation remains to be determined. In this study, we evaluated 8 different enzyme combinations (ECs) in an attempt to improve islet yield. The ECs consisted of purified, intact, or truncated class 1 (C1) and class 2 (C2) collagenases from Clostridium histolyticum (Ch) as well as neutral protease (NP) from Bacillus thermoproteolyticus rokko (thermolysin) or Ch (ChNP). Methods We report the results of 249 human islet isolations, including 99 deceased donors (research n=57, clinical n=42) and 150 chronic pancreatitis pancreases. We prepared a new enzyme mixture (NEM) composed of intact C1 and C2 collagenases and ChNP instead of using thermolysin. The NEM was first tested in split pancreas (n=5) experiments and then used for islet autologous (n=21) and allogeneic transplantation (n=10). Islet isolation outcomes from 8 different Ecs were statistically compared using multivariate analysis. Results The NEM consistently achieved higher islet yields from pancreatitis (p<0.003) and deceased donor pancreases (p<0.001) than other standard ECs. Using the NEM, islet products met release criteria for transplantation from 8 of 10 consecutive pancreases, averaging 6510±2150 IEQ/g pancreas and 694,681±147,356 total IEQ/transplantation. In autologous isolation, the NEM yielded >200,000 IEQ from 19 of 21 pancreases (averaging 422,893±181,329 total IEQ and 5979±1469 IEQ/kg recipient body weight) regardless of the severity of fibrosis. Conclusions A new enzyme mixture composed of Clostridium histolyticum neutral protease with CIzyme high intact C1 collagenase recovers higher islet yield from deceased and pancreatitis pancreases while retaining islet quality and function.

Evidence for Instant Blood-Mediated Inflammatory Reaction in Clinical Autologous Islet Transplantation

“…Previous reports have demonstrated that the insulin-independence rate after AIT is dependent on the mass of transplanted islets (13), and after intrahepatic infusion autologous islets survive better than allogenic islets (12). Despite the improvements made in islet isolation techniques (14) and the absence of an autoimmune response against the islets, achievement of insulin independence among AIT patients is rather poor at about 40% (15).…”

Human pancreatic islet transplantation: an update and description of the establishment of a pancreatic islet isolation laboratory

“…In the past 6 years, an additional obstacle was introduced into islet isolation research, as the most effective collagenase used in pancreatic digestion (Liberase HI, Roche Pharmaceutics) was removed from the market due to the discovery of an association between the enzyme and an increased risk of spongiform encephalopathy transmission (40). Other collagenases, such as Serva NB1 (Serva, Heidelberg, Germany), Liberase MTF (Roche, Indianapolis, USA) and Vitacyte (CIzyme HA, Indianapolis, USA) (37,(41)(42)(43) are promising alternatives for islet isolation (44). However, studies that have sought to assess which enzyme would be the most adequate for pancreatic digestion have produced conflicting results (40,42,(45)(46)(47)(48).…”

Different digestion enzymes used for human pancreatic islet isolation: A mixed treatment comparison (MTC) meta-analysis

“…Recently, Balamurugan et al 38 evaluated 8 different enzyme combinations in an attempt to improve islet yield. The enzyme combinations consisted of purified, intact or truncated Class I and Class II collagenases from C. histolyticum (Ch) and a NP from Bacillus thermoproteolyticus rokko (thermolysin) or from Ch.…”

“…was able to recover the highest islet yield while retaining islet quality. 38 Probably, future comparisons will be made analyzing this new enzyme mixture.…”