Derivation of xeno-free and GMP-grade human embryonic stem cells--platforms for future clinical applications

Clinically compliant human embryonic stem cells (hESCs) should be developed in adherence to ethical standards, without risk of contamination by adventitious agents. Here we developed for the first time animal-component free and good manufacturing practice (GMP)-compliant hESCs. After vendor and raw material qualification, we derived xeno-free, GMP-grade feeders from umbilical cord tissue, and utilized them within a novel, xeno-free hESC culture system. We derived and characterized three hESC lines in adherence to regulations for embryo procurement, and good tissue, manufacturing and laboratory practices. To minimize freezing and thawing, we continuously expanded the lines from initial outgrowths and samples were cryopreserved as early stocks and banks. Batch release criteria included DNA-fingerprinting and HLA-typing for identity, characterization of pluripotency-associated marker expression, proliferation, karyotyping and differentiation in-vitro and in-vivo. These hESCs may be valuable for regenerative therapy. The ethical, scientific and regulatory methodology presented here may serve for development of additional clinical-grade hESCs.     

Current regulatory issues in cell and tissue therapy

Cell-based therapies have grown dramatically in power and scope in recent years. Once limited to blood and BM transplantation, these therapies now encompass tissue repair and regeneration, metabolic support, gene replacement, and immune effector functions, with established and investigational clinical applications in disorders affecting nearly every tissue and organ system. The complexity and novel applications of human cells, tissues, and cellular and tissue-based products (HCT/Ps), however, present potential risks for adverse events. The US Food and Drug Administration, responding to these concerns, has established a tiered, risk-based regulatory structure, in which more rigorous controls and safeguards are required for products thought to pose increased risk. The proposed good tissue practices (GTP) rule and existing good manufacturing practices (GMP) requirements form the principal elements of this regulatory framework. The proposed GTPs are intended to prevent HCT/P contamination with infectious disease agents, and to ensure that these cells and tissues maintain their integrity and function. GMPs focus on production of safe, pure, and potent products, and entail a higher level of process control and product characterization. All HCT/Ps will be required to comply with GTPs. HCT/Ps considered to present greater risks of adverse events, however, will be subject to both GTPs and GMPs, and must obtain premarket approval using the Investigational New Drug (IND) mechanism established for biologics. Although these requirements will present significant challenges for clinician- investigators and laboratories producing HCT/Ps, the regulations fundamentally support good clinical care by increasing safety and control, and enable good science by improving the quality and reliability of data.     

Temporal bone bank: complying with European union directives on human tissue and cells

Background

Availability of allograft tympano-ossicular systems (ATOS) provides unique reconstructive capabilities, allowing more radical removal of middle ear pathology. To provide ATOS, the University of Antwerp Temporal Bone Bank (UATB) was established in 1988. ATOS use was stopped in many countries because of safety issues concerning human tissue transplantation. Our objective was to maintain an ATOS tissue bank complying with European Union (EU) directives on human tissues and cells.

Methods

The guidelines of the Belgian Superior Health Council, including EU directive requirements, were rigorously applied to UATB infrastructure, workflow protocols and activity. Workflow protocols were updated and an internal audit was performed to check and improve consistency with established quality systems and changing legislations. The Belgian Federal Agency of Medicines and Health Products performed an inspection to examine compliance with national legislatives and EU directives on human tissues and cells. A sample of important procedures was meticulously examined in its workflow setting next to assessment of the infrastructure and personnel.

Results

Results are reported on infrastructure, personnel, administrative workflow, procurement, preparation, processing, distribution, internal audit and inspection by the competent authority. Donors procured: 2006, 93 (45.1%); 2007, 64 (20.6%); 2008, 56 (13.1%); 2009, 79 (6.9%). The UATB was approved by the Minister of Health without critical or important shortcomings. The Ministry accords registration each time for 2?years.

Conclusions

An ATOS tissue bank complying with EU regulations on human allografts is feasible and critical to assure that the patient receives tissue, which is safe, individually checked and prepared in a suitable environment.     

Government regulation of nonhuman primate facilities

Myriad international, federal, and state laws, regulations, rules, guidelines, and standards directly affect the activities of all nonhuman primate research facilities. Federal regulations alone encompass every aspect of facility operations. They govern the procurement, possession, handling, care, and utilization of nonhuman primates, the design, construction, maintenance, and operation of the facility, and the occupational and environmental protection afforded not only facility personnel, but also the general public. Proper management of a nonhuman primate facility depends on continual monitoring of constantly changing laws and regulations applicable to the type of facility operated and research conducted. An in-house compliance assurance program is necessary to assure conformance with pertinent regulations.     

Comparison of requirements in the European Union and United States of America for pre-clinical viral safety testing of veterinary vaccines

Of paramount importance in ensuring the safety of live and inactivated veterinary vaccines is demonstration of freedom from extraneous agents in biological starting materials used in their production. Both the European Union (EU) and United States of America (US) provide regulations and guidelines on extraneous agent testing of veterinary vaccines including guidance from the Committee for Medicinal Products for Veterinary Use (CVMP), the European Pharmacopoeia (Ph. Eur.) and the USDA Code of Federal Regulations, Title 9 (9CFR). There are distinct requirements prescribed in EU and US regulations and guidelines. The differences in EU and US requirements for extraneous agent testing of starting materials are such that there may be occasions when no one test may satisfy both sets of regulations for a given scenario. For compliance with both, for global licensing purposes it may therefore be necessary to perform additional tests and/or to justify methods chosen from one set of regulations over another, based on a variety of factors.     

Exclusion of deceased donors post-procurement of tissues

The EU Tissues and Cells Directive (2004/23/EC, 2006/17/EC, 2006/86/EC) (EUTCD) provides standards for quality and safety for all aspects of banking of tissues and cells for clinical applications. Commission Directive 2006/17/EC stipulates that the complete donor record with all the medical information is assessed for suitability before releasing tissues for clinical use. The aim of this study was to investigate the medical reasons for post-procurement donor exclusion, to identify the various potential sources for gathering information about donors’ medical and behavioural history and to evaluate their contribution to maximising the safety of donations. Information was collected from the Tissue Services (TS) records of 1000 consecutive deceased donors submitted to National Health Service Blood and Transplant (NHSBT) medical officers for authorisation for release for subsequent tissue processing and then for transplantation. Of the 1000 donors 60 (6%) were excluded because they did not fulfil the donor selection requirements of the EUTCD and NHSBT donor selection guidelines. The main reasons for medical exclusion were the presence of significant local or systemic infection in 32 donors (53% of those excluded for medical reasons) and a history of past or occult malignancy in 9 donors (15% of those excluded for medical reasons) which was not identified prior to procurement. The information leading to post-procurement exclusion was obtained from autopsy reports in 35 of the 60 excluded donors for medical reasons (58%) and from the general practitioner for 10 donors (17% of those excluded for medical reasons). In summary, careful evaluation of complete donor records reduces the potential risk of disease transmission by tissue allografts and ensures compliance with regulations and guidelines. The findings may lead to changes in donor selection policies with the aim of improving efficiency without compromising safety.     

The generation of GLP-grade human embryonic stem cell banks from four clinical-grade cell lines for preclinical research

The Singapore Stem Cell Bank has generated human embryonic stem cell banks from clinical-grade cell lines ESI-017, ESI-035, ESI-049, and ESI-053. All banks were prepared and characterized according to principles of Good Laboratory Practice for quality assurance. Importantly, each cell line has clearly documented and approved ethical provenance and meets recognized standards for performance and safety. The banks are intended to facilitate the translation of stem cell research to clinical medicine by enabling early phase research and development with high-quality, low-cost cells that are also available as clinical-grade stocks.     

Derivation and propagation of human embryonic stem cell lines from frozen embryos in an animal product-free environment

The protocols described here are comprehensive instructions for deriving human embryonic stem (hES) cell lines in xeno-free conditions from cryopreserved embryos. Details are included for propagation, cryopreservation and characterization. Initial derivation is on feeder cells and is followed by adaptation to a feeder-free environment; competent technicians can perform these simplified methods easily. From derivation to cryopreservation of fully characterized initial stocks takes 3-4 months. These protocols served as the basis for standard operating procedures (SOPs), with both operational and technical components, that we set to meet good manufacturing practice (GMP) and UK regulatory body requirements for derivation of clinical-grade cells. As such, these SOPs are currently used in our current GMP compliant facility to derive hES cell lines ab initio, in an animal product-free environment; these lines are suitable for research and potentially for clinical use in cell therapy. So far, we have derived eight clinical-grade lines, which will be freely available to the scientific community after submission/accession to the UK Stem Cell Bank.     

Production, testing and perspectives of IPV and IPV combination vaccines: GSK biologicals' view.

GSK Biologicals has been involved in the production of Polio vaccine since the early start of Polio vaccination, beginning with the first generation of Inactivated Polio Vaccine (IPV). Over time, the company has developed solid industrial experience and knowledge that significantly contributes today to the quality of our Polio vaccines. GSK Biologicals' current IPV is now routinely produced according to the process defined by Van Wezel (RIVM) in the late seventies, using Vero cells and micro-carrier technology in bioreactors. In addition to compliance with current requirements (World Health Organization, European Pharmacopoeia, Code of Federal Regulations USA), the quality of the routine vaccine is guaranteed by numerous additional data related to the characterization, to the consistency, and to the validation of the process and the testing. This supplementary data package will allow, for instance, for the application of the in vitro potency testing for routine release instead of the in vivo testing. The present views on the Polio vaccine strategy for the post eradication era have portrayed a very limited role for the current IPV. The main reasons relate to post-eradication bio-containment needs and to production capacity and costs. A reevaluation of the classic approach taken to the use of the current IPV produced from wild type polio strains positions this vaccine as a real alternative to other strategies, allowing us to take advantage of the excellent performance of IPV over many years.     

Preparing clinical grade Ag-specific T cells for adoptive immunotherapy trials

The production of clinical-grade T cells for adoptive immunotherapy has evolved from the ex vivo numerical expansion of tumor-infiltrating lymphocytes to sophisticated bioengineering processes often requiring cell selection, genetic modification and other extensive tissue culture manipulations, to produce desired cells with improved therapeutic potential. Advancements in understanding the biology of lymphocyte signaling, activation, homing and sustained in vivo proliferative potential have redefined the strategies used to produce T cells suitable for clinical investigation. When combined with new technical methods in cell processing and culturing, the therapeutic potential of T cells manufactured in academic centers has improved dramatically. Paralleling these technical achievements in cell manufacturing is the development of broadly applied regulatory standards that define the requirements for the clinical implementation of cell products with ever-increasing complexity. In concert with academic facilities operating in compliance with current good manufacturing practice, the prescribing physician can now infuse T cells with a highly selected or endowed phenotype that has been uniformly manufactured according to standard operating procedures and that meets federal guidelines for quality of investigational cell products. In this review we address salient issues related to the technical, immunologic, practical and regulatory aspects of manufacturing these advanced T-cell products for clinical use.     

Proposed rule: current good manufacturing practice in manufacturing, packing, or holding dietary ingredients and dietary supplements

The Dietary Supplement Health and Education Act (DSHEA) was enacted in October 1994 to promote the health of Americans by ensuring easier access to safe dietary supplements. Many supplements such as vitamins, minerals, herbs and amino acids have been reported to be helpful in chronic conditions (i.e., heart disease, cancer and osteoporosis). Under DSHEA, dietary supplements can be marketed without prior FDA approval; the burden is on this agency to show that a marketed dietary supplement is unsafe. However, DSHEA retained the FDA's authority to issue regulations that require the manufacture of dietary supplements be in compliance with current good manufacturing practice (cGMP) standards, which are needed to ensure their quality. Several quality-related concerns of marketed dietary supplements that came to light since the passage of DSHEA prompted the FDA in 2003 to propose rules for cGMP for the manufacture, packaging and holding (storage) of dietary supplements. This review will present the highlights of these proposed rules, focusing on the legislative history of DSHEA, rationale for proposing cGMPs along with a general discussion of the specific requirements. Given the voluminous nature of the specific details, the reader is directed to the pertinent FDA publications for details. In this analysis, selected scientific and legal issues are also discussed to promote a better understanding and implications of these rules.     

Tissue banking training courses: Polish experience

Personnel directly involved in the donation, procurement, testing, processing, preservation, storage and distribution of human tissues and cells should be appropriately qualified and provided with timely and relevant training according to EU directives. In the time of new tissue and cells regulations implementation such a training system existed in Poland only at a local level. The first training programme outlines for various groups of health professionals engaged in tissue banking practice was created in co-operation with the Institute for LifeLong Learning at University of Barcelona in 2006. This initial training courses were financially supported by EU Transition Facility Programme 2004. Then, starting from 2006, based on previous experience, system of advanced training courses was created. This training programme was financially supported by the National Programme for the Development of Transplantation Medicine 2006–2009—POLGRAFT financed by Polish Ministry of Health. During 2006 and 2007 first set of tissue banking initial training courses were provided according to TF 2004 project. Over 200 pathologists, forensic medicine specialists and other medical doctors responsible for donor screening and classification, medical directors of tissue establishments, technical staff; tissue graft users: orthopaedic surgeons, neurosurgeons, cardiosurgeons and ophthalmologists were trained. Between 2006 and 2009 there were organized 8 advanced tissue banking training courses according to POLGRAFT programme. There were organized both theoretical and practical courses on various aspects of tissue for over 350 persons. We present our experience in organisation of international and national tissue banking training courses.     

Against Harmful Research on Non‐Agreeing Children

The Code of Federal Regulations permits harmful research on children who have not agreed to participate, but I will argue that it should be no more permissive of harmful research on such children than of harmful research on adults who have not agreed to participate. Of course, the Code permits harmful research on adults. Such research is not morally problematic, however, because adults must agree to participate. And, of course, the Code also permits beneficial research on children without needing their explicit agreement. This sort of research is also not problematic, this time because paternalism towards children may be justifiable. The moral problem at the center of this paper arises from the combination of two potential properties of pediatric research, first that it might be harmful and second that its subjects might not agree to participate. In Section 2 of this article I explain how the Code permits harmful research on non‐agreeing children. Section 3 contains my argument that we should no more permit harmful research on non‐agreeing children than on non‐agreeing adults. In Section 4, I argue that my thesis does not presuppose that pediatric assent has the same moral force that adult consent does. In Section 5, I argue that the distinction between non‐voluntary and involuntary research is irrelevant to my thesis. In Section 6, I rebut an objection based on the power of parental permission. In Section 7 I suggest how the Code of Federal Regulations might be changed.     

Tissue Banking in Australia

The legal structure for the regulation of tissue banking has existed for many years. In Australia, the donation of human tissue is regulated by legislation in each of the eight States and Territories. These substantially uniform Acts were passed in the late 1970's and early 1980's, based on model legislation and underpinned by the concept of consensual giving. However, it was not until the early 1990's that tissue banking came under the notice of regulatory authorities. Since then the Australian Government has moved quickly to oversee the tissue banking sector in Australia. Banked human tissue has been deemed to be a therapeutic good under the Therapeutic Goods Act 1989, and tissue banks are required to be licensed by the Therapeutic Goods Administration and are audited for compliance with the Code of Good Manufacturing Practice- Human Blood and Tissues. In addition, tissue banks must comply with a myriad of other standards, guidelines and recommendations.     

Standard Operating Procedures,ethical and legal regulations in BTB (Brain/Tissue/Bio) banking: what is still missing?

The use of human biological specimens in scientific research is the focus of current international public and professional concern and a major issue in bioethics in general. Brain/Tissue/Bio banks (BTB-banks) are a rapid developing sector; each of these banks acts locally as a steering unit for the establishment of the local Standard Operating Procedures (SOPs) and the legal regulations and ethical guidelines to be followed in the procurement and dissemination of research specimens. An appropriat Code of Conduct is crucial to a successful operation of the banks and the research application they handle. What are we still missing ? (1) Adequate funding for research BTB-banks. (2) Standard evaluation protocls for audit of BTB-bank performance. (3) Internationally accepted SOP’s which will facilitate exchange and sharing of specimens and data with the scientific community. (4) Internationally accepted Code of Conduct. In the present paper we review the most pressing organizational, methodological, medico-legal and ethical issues involved in BTB-banking; funding, auditing, procurement, management/handling, dissemination and sharing of specimens, confidentiality and data protection, genetic testing, “financial gain” and safety measures. Taking into consideration the huge variety of the specimens stored in different repositories and the enormous differences in medico-legal systems and ethics regulations in different countries it is strongly recommend that the health-care systems and institutions who host BTB-Banks will put more efforts in getting adequate funding for the infrastructure and daily activities. The BTB-banks should define evaluation protocols, SOPs and their Code of Conduct. This in turn will enable the banks to share the collected specimens and data with the largest possible number of researchers and aim at a maximal scientific spin-off and advance in public health research.     

Standard Operating Procedures,ethical and legal regulations in BTB (Brain/Tissue/Bio) banking: what is still missing?

The use of human biological specimens in scientific research is the focus of current international public and professional concern and a major issue in bioethics in general. Brain/Tissue/Bio banks (BTB-banks) are a rapid developing sector; each of these banks acts locally as a steering unit for the establishment of the local Standard Operating Procedures (SOPs) and the legal regulations and ethical guidelines to be followed in the procurement and dissemination of research specimens. An appropriat Code of Conduct is crucial to a successful operation of the banks and the research application they handle. What are we still missing ? (1) Adequate funding for research BTB-banks. (2) Standard evaluation protocls for audit of BTB-bank performance. (3) Internationally accepted SOP’s which will facilitate exchange and sharing of specimens and data with the scientific community. (4) Internationally accepted Code of Conduct. In the present paper we review the most pressing organizational, methodological, medico-legal and ethical issues involved in BTB-banking; funding, auditing, procurement, management/handling, dissemination and sharing of specimens, confidentiality and data protection, genetic testing, “financial gain” and safety measures. Taking into consideration the huge variety of the specimens stored in different repositories and the enormous differences in medico-legal systems and ethics regulations in different countries it is strongly recommend that the health-care systems and institutions who host BTB-Banks will put more efforts in getting adequate funding for the infrastructure and daily activities. The BTB-banks should define evaluation protocols, SOPs and their Code of Conduct. This in turn will enable the banks to share the collected specimens and data with the largest possible number of researchers and aim at a maximal scientific spin-off and advance in public health research.     

Program oversight enhancements (POE): the big PAM

Federal animal welfare regulations and policies require compliance during animal research. But the methods used to oversee, assess, and ascertain compliance are left in the hands of the research institution and its institutional animal care and use committee (IACUC). Differences in institutional cultures, research goals, individuals responsible for animal care and use activities and oversight, and availability of financial and personnel resources have given rise to a variety of institution-specific mechanisms for ensuring compliance with the federal requirements and specifically with IACUC-approved animal research activities. In recent years one such mechanism, postapproval monitoring (PAM), has risen in popularity. An often cited topic in animal welfare-related conferences and periodicals, it is well on its way to becoming a compliance standard. However, it is but one mechanism for ensuring postapproval compliance and is not required by the federal animal welfare regulations. In this article we describe alternative mechanisms for ensuring compliance with IACUC-approved animal research through the use of program oversight enhancements (POE) in the context of an institution's animal care and use program. We present these enhancements in a way that allows readers to pick and choose those of interest. While these methods may not be feasible at all institutions, adopting even a few should improve a program's ability to ensure compliance with approved animal research and reduce the need for an aggressive and formal postapproval monitoring process.