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Cultivated autologous limbal epithelial cell (CALEC) transplantation: Development of manufacturing process and clinical evaluation of feasibility and safety.

ABSTRACT: To treat unilateral limbal stem cell (LSC) deficiency, we developed cultivated autologous limbal epithelial cells (CALEC) using an innovative xenobiotic-free, serum-free, antibiotic-free, two-step manufacturing process for LSC isolation and expansion onto human amniotic membrane with rigorous quality control in a good manufacturing practices facility. Limbal biopsies were used to generate CALEC constructs, and final grafts were evaluated by noninvasive scanning microscopy and tested for viability and sterility. Cultivated cells maintained epithelial cell phenotype with colony-forming and proliferative capacities. Analysis of LSC biomarkers showed preservation of "stemness." After preclinical development, a phase 1 clinical trial enrolled five patients with unilateral LSC deficiency. Four of these patients received CALEC transplants, establishing preliminary feasibility. Clinical case histories are reported, with no primary safety events. On the basis of these results, a second recruitment phase of the trial was opened to provide longer term safety and efficacy data on more patients.

SUBMITTER: Jurkunas UV

PROVIDER: S-EPMC10438443 | biostudies-literature | 2023 Aug

REPOSITORIES: biostudies-literature

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Cultivated autologous limbal epithelial cell (CALEC) transplantation: Development of manufacturing process and clinical evaluation of feasibility and safety.

Jurkunas Ula V UV Yin Jia J Johns Lynette K LK Li Sanming S Negre Helene H Shaw Kit L KL Samarakoon Lassana L Ayala Allison R AR Kheirkhah Ahmad A Katikireddy Kishore K Gauthier Alex A Ong Tone Stephan S Kaufman Aaron R AR Ellender Stacey S Hernandez Rodriguez Diego E DE Daley Heather H Dana Reza R Armant Myriam M Ritz Jerome J

Science advances 20230818 33

To treat unilateral limbal stem cell (LSC) deficiency, we developed cultivated autologous limbal epithelial cells (CALEC) using an innovative xenobiotic-free, serum-free, antibiotic-free, two-step manufacturing process for LSC isolation and expansion onto human amniotic membrane with rigorous quality control in a good manufacturing practices facility. Limbal biopsies were used to generate CALEC constructs, and final grafts were evaluated by noninvasive scanning microscopy and tested for viabilit ...[more]

PMID: 37595035

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Project description:BackgroundWhile therapeutic success of the limbal tissue or cell transplantation to treat severe cases of limbal stem cell (LSC) deficiency (LSCD) strongly depends on the percentage of LSCs within the transplanted cells, prospective LSC enrichment has been hampered by the intranuclear localization of the previously reported LSC marker p63. The recent identification of the ATP-binding cassette transporter ABCB5 as a plasma membrane-spanning marker of LSCs that are capable of restoring the cornea and the development of an antibody directed against an extracellular loop of the ABCB5 molecule stimulated us to develop a novel treatment strategy based on the utilization of in vitro expanded allogeneic ABCB5+ LSCs derived from human cadaveric limbal tissue.MethodsWe developed and validated a Good Manufacturing Practice- and European Pharmacopeia-conform production and quality-control process, by which ABCB5+ LSCs are derived from human corneal rims, expanded ex vivo, isolated as homogenous cell population, and manufactured as an advanced-therapy medicinal product (ATMP). This product was tested in a preclinical study program investigating the cells' engraftment potential, biodistribution behavior, and safety.ResultsABCB5+ LSCs were reliably expanded and manufactured as an ATMP that contains comparably high percentages of cells expressing transcription factors critical for LSC stemness maintenance (p63) and corneal epithelial differentiation (PAX6). Preclinical studies confirmed local engraftment potential of the cells and gave no signals of toxicity and tumorgenicity. These findings were sufficient for the product to be approved by the German Paul Ehrlich Institute and the U.S. Food & Drug Administration to be tested in an international multicenter phase I/IIa clinical trial (NCT03549299) to evaluate the safety and therapeutic efficacy in patients with LSCD.ConclusionBuilding upon these data in conjunction with the previously shown cornea-restoring capacity of human ABCB5+ LSCs in animal models of LSCD, we provide an advanced allogeneic LSC-based treatment strategy that shows promise for replenishment of the patient's LSC pool, recreation of a functional barrier against invading conjunctival cells and restoration of a transparent, avascular cornea.

Project description:BackgroundAccess to exercise for cancer survivors is poor despite global recognition of its benefits. Telerehabilitation may overcome barriers to exercise for cancer survivors but is not routinely offered.ObjectiveFollowing the rapid implementation of an exercise-based telerehabilitation program in response to COVID-19, a process evaluation was conducted to understand the impact on patients, staff, and the health service with the aim of informing future program development.MethodsA mixed methods evaluation was completed for a telerehabilitation program for cancer survivors admitted between March and December 2020. Interviews were conducted with patients and staff involved in implementation. Routinely collected hospital data (adverse events, referrals, admissions, wait time, attendance, physical activity, and quality of life) were also assessed. Patients received an 8-week telerehabilitation intervention including one-on-one health coaching via telehealth, online group exercise and education, information portal, and home exercise prescription. Quantitative data were reported descriptively, and qualitative interview data were coded and mapped to the Proctor model for implementation research.ResultsThe telerehabilitation program received 175 new referrals over 8 months. Of those eligible, 123 of 150 (82%) commenced the study. There were no major adverse events. Adherence to health coaching was high (674/843, 80% of scheduled sessions), but participation in online group exercise classes was low (n=36, 29%). Patients improved their self-reported physical activity levels by a median of 110 minutes per week (IQR 90-401) by program completion. Patients were satisfied with telerehabilitation, but clinicians reported a mixed experience of pride in rapid care delivery contrasting with loss of personal connections. The average health service cost per patient was Aus $1104 (US $790).ConclusionsTelerehabilitation is safe, feasible, and improved outcomes for cancer survivors. Learnings from this study may inform the ongoing implementation of cancer telerehabilitation.

Project description:BackgroundAutologous cultivated oral mucosal epithelial transplantation (COMET) is an important treatment for limbal stem cell deficiency. However, peripheral corneal neovascularization after surgery hinders its application. This study aims to employ a culture system using allogenic limbal niche cells (LNCs) instead of mouse-derived 3T3 cells as a feeder layer that could relieve postoperative neovascularization.MethodsRat oral mucosal epithelial cells (OMECs) were co-cultured with rat LNCs or 3T3 cells. Cultivated oral mucosal epithelial cells (COMECs) of different culture systems were identified by hematoxylin and eosin staining and immunocytochemistry. The expression levels of the angiogenesis-related factors were analyzed by RT-qPCR and western blotting/ELISA. Angiogenic potential was reconfirmed by cell viability and tube formation assays with human umbilical vein endothelial cells (HUVECs).ResultsCOMECs were obtained from both culture systems successfully. Immunocytochemistry showed approximately equal percentages of positive staining cells for p63α (p = 0.9177), ABCG2 (p = 0.526), Ki67 (p = 0.0987), and CK3 (p = 0.4000) in COMECs of different groups. RT-qPCR and western blotting/ELISA showed that COMECs of the LNC group expressed a significantly lower amount of basic fibroblast growth factor (bFGF) (p = 0.0038 for RT-qPCR, p = 0.0026 for western blotting) but more pigment epithelium-derived factor (PEDF) (p = 0.0172 for RT-qPCR, p = 0.0253 for western blotting) and soluble fms-like tyrosine kinase-1 (sFlt-1) (p < 0.0001 for RT-qPCR, p = 0.0064 for ELISA) than the COMECs of the 3T3 group. Furthermore, compared with COMECs of the 3T3 group, COMECs of the LNC group could reduce the viability (p = 0.0002) and tube formation (p = 0.0002) of HUVECs.ConclusionsLNCs could substitute 3T3 cells for expanding OMECs in vitro, and the COMECs obtained in this system are less likely to induce postsurgical neovascularization, which provides an alternative option for an ex vivo culture system and promotes the application of COMET.

Dataset Information

Cultivated autologous limbal epithelial cell (CALEC) transplantation: Development of manufacturing process and clinical evaluation of feasibility and safety.

Publications

Cultivated autologous limbal epithelial cell (CALEC) transplantation: Development of manufacturing process and clinical evaluation of feasibility and safety.

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