Patent ID: 9493743
Filing Date: 2016-11-15
CPC Classification: A01K,C12N,G01N

Claim Text:
1. A method of preparing human pancreatic beta cells or human beta cell tumors, wherein said human pancreatic beta cells or human beta cell tumors express insulin and PDX-1, said method comprising the steps of: a) dissociating neonatal human pancreatic tissue with collagenase in order to obtain neonatal human pancreas cells, b) transducing and co-transducing the neonatal human pancreas cells obtained in step a) with i) a lentiviral vector expressing only SV40 LargeT antigen under the control of the insulin promoter or ii) with a lentiviral vector expressing only SV40 LargeT antigen under the control of the insulin promoter and a lentiviral vector expressing only hTERT under the control of the insulin promoter, or iii) a lentiviral vector expressing both SV40 LargeT antigen and hTERT but no other oncogene, c) introducing the transduced neonatal pancreas obtained in b) into the kidney capsule of a first severe combined immunodeficiency (scid) mouse, d) allowing the transduced pancreas cells to develop insulinoma-like structures, wherein the neonatal human pancreas cells in insulinoma-like structures have differentiated in insulin-producing pancreatic beta cells; e) micro-dissecting the insulinoma-like structures obtained in step d), dissociating the cells thereof, f) sub-transplanting the cells obtained in step e) into the kidney capsule of a second scid mouse, g) allowing the sub-transplanted cells in step f) to develop and regenerate insulinoma-like structures, wherein said newly developed insulinoma-like structures are enriched in insulin producing pancreatic beta cells; h) micro-dissecting the insulinoma-like structures obtained in step g), and dissociating and collecting the cells thereof, i) optionally, sub-transplantating the cells obtained in step g) into the kidney capsule of a third scid mouse, to further enrich and amplify insulin-producing pancreatic beta cells; and j) optionally repeating steps f), g) and h) until an appropriate amount of insulin-producing pancreatic beta cells is obtained, wherein the insulin-producing pancreatic beta cells allow insulinoma formation and restoration of normoglycemia in diabetic SCID mice.