Prospects for the manufacture of red cells for transfusion

Whilst red cell transfusion is a well established cellular therapy, the problems of insufficiency of supply, transfusion transmitted infections and the requirement for immunological matching persist. The possibility of generating large numbers of O Rh D negative red cells at Good Manufacturing Practice grade as a route to circumvent these issues is therefore an attractive proposition. Significant numbers of erythrocytes can be generated from somatic haematopoietic stem cells, but it seems unlikely that these can provide sufficient volumes for large scale manufacture. However, human embryonic stem cells (hESC) and, potentially, induced pluripotent stem cells (iPSC), may provide a route to this objective. Red cell transfusion is an attractive goal for pluripotent stem cell‐derived therapeutics because it is a well‐characterised single cell suspension that lacks nucleated cells and has a low expression of human leucocyte antigen molecules, but many challenges remain in translating this cellular therapy to the clinic.     

Cell therapies and regenerative medicine

Molecular and cell biology has resulted in major advances in our understanding of disease pathogenesis as well as in novel strategies for the diagnosis, therapy and prevention of human diseases. Based on modern molecular, genetic and biochemical methodologies, it is on the one hand possible to identify disease-related point mutations and single nucleotide polymorphisms, for example. On the other hand, using high throughput array and other technologies, it is for example possible to simultaneously analyze thousands of genes or gene products (RNA and proteins), resulting in an individual gene or gene expression profile (‘signature’). Such data increasingly allow defining the individual disposition for a given disease and predicting disease prognosis as well as the efficacy of therapeutic strategies in the individual patient (‘personalized medicine’). At the same time, the basic discoveries in cell biology, including embryonic and adult stem cells, induced pluripotent stem cells, genetically modified cells and others, have moved regenerative medicine into the center of biomedical research worldwide with a major translational impact on tissue engineering as well as transplantation medicine. All these aspects have greatly contributed to the recent advances in regenerative medicine and the development of novel concepts for the treatment of many human diseases, including liver diseases.     

Experts’ considerations on HLA‐haploidentical stem cell transplantation

Recently, novel strategies to control graft‐versus‐host disease and facilitate engraftment have allowed an increasing number of human leukocyte antigen (HLA)‐haploidentical hematopoietic stem cell transplantation (haploHSCT) to be performed. A meeting was convened to review the biological rationale and the clinical results of various T‐cell‐depleted (TCD) and T‐cell‐replete (TCR) HLA‐haploidentical ‘transplant platforms’. The objective of the meeting was to promote discussion and consent among leading researchers in the field on three main crucial issues for haploHSCT: (i) eligibility criteria, (ii) choice of the most suitable donor, and (iii) choice of the most appropriate transplant platform. The experts in attendance agreed that a patient who is eligible for an allogeneic transplant and lacks an HLA‐identical sibling or an HLA‐matched unrelated donor should be considered for an alternative donor transplant. Together with the experience of the individual center, the most important decision criteria in choosing an alternative donor source should be the rapidity of transplantation so as to avoid disease relapse/progression. The choice of the mismatched donor should be driven by younger age, ABO blood group compatibility, and Cytomegalovirus status. If a TCD transplant is planned, NK‐alloreactive donors and/or the mother should be preferred. Prospective comparative studies are needed to establish the relative efficacy of different transplant platforms. However, expertise in stem cell manipulation and in adoptive immunotherapy is essential if a TCD transplant platform is chosen.     

CAR T in adult ALL: When and for whom?

Chimeric antigen receptor T cell therapy targeting CD19 (CART19) has shown remarkable results in patients with relapsed/refractory (r/r) B cell acute lymphoblastic leukemia (ALL). In patients 25 years of age or younger CART19 therapy is an accepted standard of care, while the treatment of older adults is less straight forward and possible only in the context of a clinical trial. Treatment of older patients with CAR T cells requires careful consideration of overall treatment goals, suitability of a consolidative hematopoietic stem cell transplant (HSCT), alternative treatment options, patient risk profile, and anticipated responses and toxicities of the specific CAR T cell products available. Here we use patient guided examples to inform approaches to care.     

Infection prevention strategies in a stem cell transplant unit: impact of change of care in isolation practice and routine use of high dose intravenous immunoglobulins on infectious complications and transplant related mortality

Objective: Nursing in ‘live islands’ and routine high dose intravenous immunoglobulins after allogeneic hematopoietic stem cell transplantation were abandoned by many teams in view of limited evidence and high costs. Methods: This retrospective single‐center study examines the impact of change from nursing in ‘live islands’ to care in single rooms (SR) and from high dose to targeted intravenous immunoglobulins ( IVIG) on mortality and infection rate of adult patients receiving an allogeneic stem cell or bone marrow transplantation in two steps and three time cohorts (1993–1997, 1997–2000, 2000–2003). Results: Two hundred forty‐eight allogeneic hematopoetic stem cell transplantations were performed in 227 patients. Patient characteristics were comparable in the three cohorts for gender, median age, underlying disease, and disease stage, prophylaxis for graft versus host disease ( GvHD) and cytomegalovirus constellation. The incidence of infections (78.4%) and infection rates remained stable (rates/1000 days of neutropenia for sepsis 17.61, for pneumonia 6.76). Cumulative incidence of GvHD and transplant‐related mortality did not change over time. Conclusions: Change from nursing in ‘live islands’ to SR and reduction of high dose to targeted IVIG did not result in increased infection rates or mortality despite an increase in patient age. These results support the current practice.     

Advances in understanding the leukaemia microenvironment

Dynamic interactions between leukaemic cells and cells of the bone marrow are a feature of haematological malignancies. Two distinct microenvironmental niches in the bone marrow, the ‘osteoblastic (endosteal)’ and ‘vascular’ niches, provide a sanctuary for subpopulations of leukaemic cells to evade chemotherapy‐induced death and allow acquisition of drug resistance. Key components of the bone marrow microenvironment as a home for normal haematopoietic stem cells and the leukaemia stem cell niches, and the molecular pathways critical for microenvironment/leukaemia interactions via cytokines, chemokines and adhesion molecules as well as hypoxic conditions, are described in this review. Finally, the genetic abnormalities of leukaemia‐associated stroma are discussed. Further understanding of the contribution of the bone marrow niche to the process of leukaemogenesis may provide new targets that allow destruction of leukaemia stem cells without adversely affecting normal stem cell self‐renewal.     

Precision medicines for B‐cell leukaemias and lymphomas; progress and potential pitfalls

There is now a plethora of new precision medicines for B‐cell malignancy including ‘classical’ kinase inhibitors, rationally designed inhibitors of anti‐apoptotic proteins and antibody or antibody drug/toxin conjugates with functional properties. Some are showing spectacular single agent activity in early phase clinical studies and may reduce or, in combination, even obviate the need for chemotherapy. Nevertheless, significant problems remain if these medicines are to be introduced into routine clinical practice in a rational and affordable manner. Firstly, precision medicines must be carefully matched in a mechanistic fashion with specific subtypes of disease. Whilst sensitivity may be predicted by the detection of key mutations or by expression of target molecules, for therapies that depend on intact intracellular signalling pathways, functional assessment on viable primary malignant cells will be necessary using assays that faithfully mimic in vivo conditions. A second, but no less important challenge is to define mechanism‐based synergistic combinations associated with minimal toxicities rather than simply adding new precision medicines to existing chemotherapeutic regimens. Finally, a closer, open, two‐way interaction between academic medicine and the pharmaceutical industry will be necessary to achieve these aims. Implementing such changes would change radically how and where patients with B‐cell malignancies are managed.     

Stem cell-based regenerative opportunities for the liver: State of the art and beyond

The existing mismatch between the great demand for liver transplants and the number of available donor organs highlights the urgent need for alternative therapeutic strategies in patients with acute or chronic liver failure. The rapidly growing knowledge on stem cell biology and the intrinsic repair processes of the liver has opened new avenues for using stem cells as a cell therapy platform in regenerative medicine for hepatic diseases. An impressive number of cell types have been investigated as sources of liver regeneration: adult and fetal liver hepatocytes,intrahepatic stem cell populations,annex stem cells,adult bone marrow-derived hematopoietic stem cells,endothelial progenitor cells,mesenchymal stromal cells,embryonic stem cells,and induced pluripotent stem cells. All these highly different cell types,used either as cell suspensions or,in combination with biomaterials as implantable liver tissue constructs,have generated great promise for liver regeneration. However,fundamental questions still need to be addressed and critical hurdles to be overcome before liver cell therapy emerges. In this review,we summarize the state-of-the-art in the field of stem cell-based therapies for the liver along with existing challenges and future perspectives towards a successful liver cell therapy that will ultimately deliver its demanding goals.     

Multiple myeloma: chemotherapy or transplantation in the era of new drugs

Objective: To review the current results of studies incorporating novel agents in multiple myeloma (MM) and discuss the role of autologous stem‐cell transplantation (ASCT) in the era of new active drugs for the treatment of this disease. The outlook for patients with symptomatic MM is changing with the introduction of bortezomib, thalidomide, and lenalidomide into the repertoire of available chemotherapeutic agents. Compared with standard chemotherapy, a survival benefit has been reported for the first time in 30 yrs. Methods: Articles published in English between 1969 and 2008 were identified by searching PubMed for ‘myeloma’, ‘diagnosis’, ‘thalidomide’, ‘bortezomib’, ‘lenalidomide’, ‘dexamethasone’, ‘prednisone’, ‘doxorubicin’, ‘cyclophosphamide’, ‘melphalan’, ‘combination chemotherapy’, and ‘autologous transplantation’. Results: In randomized studies, bortezomib, thalidomide, and lenalidomide have each been combined with dexamethasone, alkylating agents, or doxorubicin, and such combinations resulted in significant improvement in progression‐free survival. Conclusions: The incorporation of new drugs as induction therapy along with ASCT appears to produce very good partial response rates, slightly superior to those achieved by conventional chemotherapy with new drugs. How to best optimize induction, consolidation, and maintenance therapy and how to best select and prepare patients for ASCT are still to be determined. Randomized trials are needed to directly compare the current best chemotherapeutic approach with best ASCT strategies and to guide clinical practice for patients with MM.     

Clinical grade cell manipulation

In the past decade, a new form of therapy based on biological rather than pharmacological intervention has been developed. The term 'cell therapy', as applied to this new therapeutic tool, means the administration of living, non-germline somatic cells to humans for diagnostic or therapeutic purposes. Cell therapy products (CTPs) are generated by ex vivo processes, which comprise cell harvesting from patients or healthy donors, in vitro manipulation and administration of the manipulated cells to patients. The aim of ex vivo processes is to obtain cell subsets with defined functional properties that are capable of replacing or repairing damaged tissues or organs. Some examples of cell therapy are transplantation of expanded haematopoietic stem cells (HSCs), adoptive immunotherapy and dendritic cell vaccination to augment or restore the immune response for the treatment of malignant or infectious diseases. The types of cells most frequently used for cell therapy include haematopoietic pluripotent progenitor and stem cells from the bone marrow and peripheral blood, T-cell clones and dendritic cells. Although CTPs should be produced according to good manufacturing practice, they differ from traditional pharmaceutical products with regard to quality control and safety aspects. These differences prompted the development of a number of documents issued by regulatory bodies, which specifically address CTPs. This review discusses several issues related to the design, construction and validation of a hospital-based facility for the production of CTPs, the implementation of cell-manipulation processes and quality control of the final products.     

A case‐based discussion of clinical problems in the management of patients treated with ruxolitinib for myelofibrosis

Ruxolitinib is a dual janus kinase 1 (JAK1)/JAK2 inhibitor used to treat splenomegaly and symptoms associated with myelofibrosis (MF). Current therapeutic options for symptomatic MF include supportive care, myelosuppressive therapy (such as hydroxycarbamide) and janus kinase (JAK) inhibitors (in particular ruxolitinib). Allogeneic stem cell transplantation remains the only potentially curative treatment for MF, and younger transplant‐eligible patients should still be considered for allogeneic stem cell transplantation; however, this is applicable only to a small proportion of patients. There is now increasing and extensive experience of the efficacy and safety of ruxolitinib in MF, both in clinical trials and in ‘real‐world’ practice. The drug has been shown to be of benefit in intermediate‐1 risk patients with symptomatic splenomegaly or other MF‐related symptoms, and higher risk disease. Optimal use of the drug is required to maximise clinical benefit, requiring an understanding of the balance between dose‐dependent responses and dose‐limiting toxicities. There is also increasing experience in the use of ruxolitinib in the pre‐transplantation setting. This paper aims to utilise several ‘real‐life’ cases to illustrate several strategies that may help to optimise clinical practice.     

10 years of intracoronary and intramyocardial bone marrow stem cell therapy of the heart: from the methodological origin to clinical practice

Intracoronary and intramyocardial stem cell therapy aim at the repair of compromised myocardium thereby--as a causal treatment--preventing ventricular remodeling and improving overall performance. Since the first-in-human use of bone marrow stem cells (BMCs) after acute myocardial infarction in 2001, a large number of clinical studies have demonstrated their clinical benefit: BMC therapy can be performed with usual cardiac catheterization techniques in the conscious patient as well as also easily during cardiosurgical interventions. New York Heart Association severity degree of patients as well as physical activity improve in addition to ("on top" of) all other therapeutic regimens. Stem cell therapy also represents an ultimate approach in advanced cardiac failure. For acute myocardial infarction and chronic ischemia, long-term mortality after 1 and 5 years, respectively, is significantly reduced. A few studies also indicate beneficial effects for chronic dilated cardiomyopathy. The clinical use of autologous BMC therapy implies no ethical problems, when unmodified primary cells are used. With the use of primary BMCs, there are no major stem cell-related side effects, especially no cardiac arrhythmias and inflammation. Various mechanisms of the stem cell action in the human heart are discussed, for example, cell transdifferentiation, cell fusion, activation of intrinsic cardiac stem cells, and cytokine-mediated effects. New techniques allow point-of-care cell preparations, for example, within the cardiac intervention or operation theater, thereby providing short preparation time, facilitated logistics of cell transport, and reasonable cost effectiveness of the whole procedure. The 3 main indications are acute infarction, chronic ischemic heart failure, and dilated cardiomyopathy. Future studies are desirable to further elucidate the mechanisms of stem cell action and to extend the current use of intracoronary and/or intramyocardial stem cell therapy by larger and presumably multicenter and randomized trials.     

Sirolimus for the treatment of multi-resistant pure red cell aplasia

Patients with relapsed, refractory or advanced stage B non-Hodgkin lymphoma (NHL) continue to have a dismal prognosis. This review summarises current and novel cellular and immunotherapy for these high-risk populations, including haematopoietic stem cell transplant, bispecific antibodies, viral-derived cytotoxic T cells, chimeric antigen receptor (CAR) T cells, and natural killer (NK) cell therapy, as discussed at the 6th International Symposium on Childhood, Adolescent and Young Adult Non-Hodgkin Lymphoma on September 26th–29th 2018 in Rotterdam, the Netherlands, and explores the future of NK/CAR NK therapies.     

Update on hepatic stem cells

Abstract: The liver, like most organs in an adult healthy body, maintains a perfect balance between cell gain and cell loss. Though normally proliferatively quiescent, simple hepatocyte loss such as that caused by partial hepatectomy, uncomplicated by virus infection or inflammation, invokes a rapid regenerative response to restore liver mass. This restoration of moderate cell loss and ‘wear and tear’ renewal is largely achieved by hepatocyte self‐replication. Furthermore, cell transplant models have shown that hepatocytes can undergo significant clonal expansion. Such observations indicate that hepatocytes are the functional stem cells of the liver. More severe liver injury activates a facultative stem cell compartment located within the intrahepatic biliary tree, giving rise to cords of biliary epithelia within the lobules before these cells differentiate into hepatocytes. A third population of stem cells with hepatic potential resides in the bone marrow; these haematopoietic stem cells can contribute to the albeit low renewal rate of hepatocytes, make a more significant contribution to regeneration, and even completely restore normal function in a murine model of hereditary tyrosinaemia. How these three stem cell populations integrate to achieve a homeostatic balance is not understood. This review focuses on three aspects of liver stem cell biology: 1) the hepatic stem cell candidates; 2) models of cell transplantation into the liver; and 3) the therapeutic potential of hepatic stem cells.     

Cancer stem cells in lung cancer: Evidence and controversies

The cancer stem cell (CSC) model is based on a myriad of experimental and clinical observations suggesting that the malignant phenotype is sustained by a subset of cells characterized by the capacity for self‐renewal, differentiation and innate resistance to chemotherapy and radiation. CSC may be responsible for disease recurrence after definitive therapy and may therefore be functionally synonymous with minimal residual disease. Similar to other solid tumours, several putative surface markers for lung CSC have been identified, including CD133 and CD44. In addition, expression and/or activity of the cytoplasmic enzyme aldehyde dehydrogenase ALDH and capacity of cells to exclude membrane permeable dyes (known as the ‘side population’) correlate with stem‐like function in vitro and in vivo. Embryonic stem cell pathways such as Hedgehog, Notch and WNT may also be active in lung cancers stem cells and therefore may be therapeutically targetable for maintenance therapy in patients achieving a complete response to surgery, radiotherapy or chemotherapy. This paper will review the evidence regarding the existence and function of lung CSC in the context of the experimental and clinical evidence and discuss some ongoing controversies regarding this model.     

Haemophilia management: time to get personal?

Summary. The possibility of alloimmunization in patients receiving protein replacement therapy depends on (at least) three risk factors, which are necessary concomitantly but insufficient alone. The first is the degree of structural difference between the therapeutic protein and the patient’s own endogenous protein, if expressed. Such differences depend on the nature of the disease mutation and the pre‐mutation endogenous protein structure as well as on post‐translational changes and sequence‐engineered alterations in the therapeutic protein. Genetic variations in the recipients’ immune systems comprise the second set of risk determinants for deleterious immune responses. For example, the limited repertoire of MHC class II isomers encoded by a given person’s collection of HLA genes may or may not be able to present a ‘foreign’ peptide(s) produced from the therapeutic protein – following its internalization and proteolytic processing – on the surface of their antigen‐presenting cells (APCs). The third (and least characterized) variable is the presence or absence of immunologic ‘danger signals’ during the display of foreign‐peptide/MHC‐complexes on APCs. A choice between existing therapeutic products or the manufacture of new proteins, which may be less immunogenic in some patients or patient populations, may require prior definition of the first two of these variables. This leads then to the possibility of developing personalized therapies for disorders due to genetic deficiencies in endogenous proteins, such as haemophilia A and B. [Correction made after online publication 11 July 2011: several critical corrections have been made to the abstract].     

Addiction medicine ethics: relapse,no lapse and the struggle to treat addicts like everyone else

Two case studies are presented as a focus for discussion of ethics in addiction medicine. The first is that of the alcohol‐dependent patient who receives a liver transplant. The second is that of a heroin‐dependent patient who continues to inject himself while in a general medical ward. I make some comments about the obligations of doctors to treat those who cause harm to themselves as they would treat those who are ‘not responsible’.     

Mesenchymal stem cells in the treatment of chronic lung disease

Mesenchymal stem cells (MSCs) are a populace of non‐haematopoietic multipotent stromal cells, which have the ability to differentiate into tissue derived from a single germ layer. MSCs have been isolated from various sites, including adipose tissue, skeletal muscle, synovium, spleen, thymus, lung and amniotic fluid, but are most often isolated from bone marrow. MSCs have several valuable functions that make them a promising therapeutic option in the field of regenerative medicine, including the secretion of anti‐inflammatory cytokines and growth factors, the migration of cells to the site of injury when administered and the ability to ‘rescue’ cells through the transfer of functional mitochondria. They also offer the possibility of autologous cell transplantation, circumventing immune rejection. These properties, among others, make MSCs a promising potential therapeutic agent in the treatment of chronic lung diseases with high rates of morbidity and mortality, such as idiopathic pulmonary fibrosis (IPF), COPD and obstructive bronchiolitis (OB). Numerous animal models have shown the protective and reparative effects of MSCs in models of experimental lung injury. There are currently several clinical trials underway to evaluate the safety and efficacy of MSCs in the treatment of IPF, COPD and OB. While early results are encouraging, a considerable amount of research must be done concerning the safety MSCs, as well as their optimal dosage, time and route of administration. In addition, much is still unknown about the pathogenesis of these chronic lung diseases, as well as the mechanisms MSCs utilize to assist in their repair.     

Isolated cerebral manifestation of Epstein–Barr virus‐associated post‐transplant lymphoproliferative disorder after allogeneic hematopoietic stem cell transplantation: a case of clinical and diagnostic challenges

N.A. Kittan, F. Beier, K. Kurz, H.H. Niller, L. Egger, W. Jilg, R. Andreesen, E. Holler, G.C. Hildebrandt. Isolated cerebral manifestation of Epstein–Barr virus‐associated post‐transplant lymphoproliferative disorder after allogeneic hematopoietic stem cell transplantation: a case of clinical and diagnostic challenges.
Transpl Infect Dis 2011: 13: 524–530. All rights reserved Abstract: We present the case of a 49‐year‐old male patient with Epstein–Barr virus (EBV)‐associated post‐transplant lymphoproliferative disorder (PTLD) limited to the brain that occurred 6 months after allogeneic hematopoietic stem cell transplantation (HSCT). Clinical symptoms included mental confusion, ataxia, and diplopia. Magnetic resonance imaging (MRI) revealed cerebellar and periventricular lesions consistent with an inflammatory process. Cerebrospinal fluid (CSF) analysis, but not peripheral blood, was positive for EBV‐DNA, but no malignant cells were found. Brain biopsy was not feasible because of low platelet counts. As we considered a diagnosis of either EBV‐associated encephalitis or PTLD, the patient was treated with rituximab combined with antiviral therapy. However, the cerebral lesions progressed and follow‐up CSF testing revealed immunoglobulin H clonality as evidence of a malignant process. Subsequent treatment attempts included 2 donor lymphocyte infusions (DLI). Despite treatment, the patient died from autopsy‐proven PTLD within 8 weeks of the onset of symptoms. This case demonstrates the clinical and diagnostic challenges of primary cerebral PTLD in a patient following allogeneic HSCT.     

Sequential liver and haematopoietic stem cell transplantation in a case of fulminant hepatitis associated liver failure and aplastic anaemia

The management of severe aplastic anaemia is particularly challenging when it occurs in the context of recent liver transplantation. Rapid identification of a suitable donor followed by allogeneic haematopoietic stem cell transplantation is the only curative option. This scenario is often complicated by potentially life‐threatening infections that develop as a consequence of immunosuppression. Alternative donor transplantation using suitably matched unrelated donors can be potentially life‐saving when suitably matched sibling donors are unavailable. Above all, a dedicated interdisciplinary approach with seamless communication between hepatology, transplant surgery, haematology, and stem cell transplant services is essential to achieving optimal outcomes. Herein, we describe a case of severe hepatitis leading to hepatic failure who was treated with liver transplantation from a deceased donor, and later received an allogeneic haematopoietic stem cell transplantation from a matched unrelated donor for hepatitis‐associated aplastic anaemia.