A phase I study of MAGE-A1-targeted T1367 T-cell receptor-based cell therapy in patients with advanced multiple myeloma.

Not available.

Long-Term Follow-Up After the Application of Mesenchymal Stromal Cells in Children and Adolescents with Steroid-Refractory Graft-Versus-Host Disease.

Steroid-refractory graft-versus-host disease (GvHD) is a life-threatening complication after allogeneic hematopoietic stem cell transplantation (alloHSCT). Alternative treatment options are often insufficient. Several studies have proven the efficacy of mesenchymal stromal cells (MSCs) in the treatment of therapy-refractory acute GvHD in adult and pediatric patients. Long-term data in pediatric patients are scarce. In this retrospective analysis, a total of 25 patients with a median age of 10.6 years (range 0.6-22.1 years) who received bone marrow-derived MSCs after alloHSCT for the treatment of steroid-refractory III and IV GvHD were analyzed. The median observation period of the surviving patients was 9.3 years (1.3-12.7 years) after HSCT. Among the 25 patients, 10 (40.0%) died [relapse (n = 3), multiorgan failure (n = 6), cardiorespiratory failure (n = 1)] at median 0.5 years (0.2-2.3 years) after HSCT. Partial response and complete remission (PR, CR) of the GvHD were achieved in 76.0% and 24.0% of the patients, respectively. Transplant-related mortality was 0% in the patients who achieved CR after MSC treatment and 26.3% for those with PR. A median improvement by one intestinal or liver GvHD stage (range 1-4) could be achieved after MSC application. No potentially MSC-related long-term adverse effects, for example, secondary malignancy, were identified. In conclusion, the intravenous application of allogeneic MSCs was safe and proved effective for the treatment of steroid-refractory GvHD. However, larger, prospective, and randomized trials are needed to evaluate these findings.

Longtime Outcome After Intraosseous Application of Autologous Mesenchymal Stromal Cells in Pediatric Patients and Young Adults with Avascular Necrosis After Steroid or Chemotherapy.

Avascular necrosis (AVN) is a severe complication of immunosuppressant therapy or chemotherapy. A beneficial AVN therapy with core decompression (CD) and intraosseous infusion of mesenchymal stromal cells (MSCs) has been described in adult patients, but there are only few data on MSC applications in pediatric and young adult patients (PYAP). Between 2006 and 2015, 14 AVN lesions of 10 PYAP (6 females) with a median age of 16.9 years (range 8.5-25.8 years) received CD and intraosseous application of autologous MSCs. Data of these patients were analyzed regarding efficacy, safety, and feasibility of this procedure as AVN therapy and compared with a control group of 13 AVN lesions of 11 PYAP (5 females) with a median age of 17.9 years (range 13.5-27.5 years) who received CD only. During the follow-up analysis [MSC group: median 3.1 (1.6-5.8) years after CD; CD group: median 2.0 (1.5-8.5) years after CD], relative lesion sizes (as assessed by magnetic resonance imaging) compared with the initial lesion volume, were significantly lower (P < 0.05) in the MSC group (volume reduction to a median of 18.5%) when compared with the CD group (58.0%). One lesion in the MSC group comprised a complete remission. Size progression was not observed in either group. Clinical improvement (pain, mobility) was not significantly different between the two groups. None of the patients experienced treatment-related adverse effects. CD and additional MSC application was regarded safe, effective, feasible, and superior in reducing the lesion size when compared with CD only. Prospective, randomized clinical trials are needed to further evaluate these findings.

Collagen cell carriers seeded with human urothelial cells for urethral reconstructive surgery: first results in a xenograft minipig model.

PURPOSE: Urethral strictures are a common disease of the lower urinary tract in men. At present, the use of buccal mucosa is the method of choice for long or recurrent strictures. However, autologous tissue-engineered grafts are still under investigation for reconstructive urological surgery. The aim of this pilot study was to evaluate the use of human urothelial cells (HUC) seeded on bovine collagen type I-based cell carriers (CCC) in an animal model and to evaluate short-term outcome of the surgical procedure. METHODS: Four male Göttingen minipigs were used with immunosuppression (cyclosporine A) for this pilot xenograft study. HUC obtained from human benign ureteral tissue were stained by PKH26 and seeded on a collagen cell carrier (CCC). Seven weeks after urethral stricture induction and protective vesicostomy, cell-seeded CCC was implanted in the urethra with HUC luminal and antiluminal, respectively. After two weeks animals were euthanized, urethrography and histological assessment were performed. RESULTS: Surgery was technically feasible in all minipigs. Stricture was radiologically established 7 weeks after induction. CCC was visible after two weeks and showed good integration without signs of inflammation or rejection. In the final urethrography, no remaining stricture could be detected. Near porcine urothelium, PKH26-positive areas were found even if partially detached from CCC. Although diminished, immunofluorescence with pankeratin, CK20, E-cadherin and ZO-1 showed intact urothelium in several areas on and nearby CCC. CONCLUSION: Finally, this study demonstrates that the HUC-seeded CCC used as a xenograft in minipigs is technically feasible and shows promising results for further studies.

Precise injection of human mesenchymal stromal cells in the urethral sphincter complex of Göttingen minipigs without unspecific bulking effects.

AIM: To investigate if injection of cells in the urethral sphincter complex causes unspecific bulking effects. METHODS: Human mesenchymal stromal cells were isolated, expanded, and characterized. For transurethral injection, cells were labeled with the fluorescent dye PKH26 and in magnetic resonance imaging associated experiments with superparamagnetic particles. Aliquots of cells in 250 µL solvent were injected under vision in the urethral sphincter of immuno-suppressed Göttingen minipigs. Sphincteric closure pressure was recorded by standard and high-definition urethral pressure profilometry prior to and after cell injection. The animals were sacrificed after surgery or after 3 weeks, 3, 6, or 12 months of follow-up. The localisation of the injected cells was explored by histochemistry. Sham-treated animals served as controls. RESULTS: PKH26-labeled cells survive injections in sphincter tissue samples by Williams cystoscopic injection needle well. In our animal study, the cellular depots were detected in the submucosa or in deeper zones of the sphincter, depending of the length of the injection needle (4-8 mm). Adverse effects associated with injection of cells or solvent such as a noteworthy bleeding, incontinence, or obstruction, were not recorded (n = 96 minipigs). However, a transient infiltration of macrophages was detected 3 weeks after cell injection. Changes in the urethral pressure profiles were not observed in cell-treated (n = 72) compared to sham-treated animals (n = 24). CONCLUSIONS: Injection of small aliquots of cells to investigate cell therapies in minipigs is a feasible and safe procedure, and it does not bias the intrinsic urethral wall pressure.

Hypothermia Promotes Survival of Ischemic Retinal Ganglion Cells.

PURPOSE: Ischemic stroke in retinal arteries leads to death of neural tissue and ultimately to blindness. The retina is known to die within 4 hours after onset of ischemia. It is debated whether hypothermia might increase the time window for medical treatment and thereby the chance of recovering sight. In order to characterize the time course of cell death during ischemia and potential beneficial effects of hypothermia in more detail, we investigated the survival of ganglion cells in ischemic pig and human retina as a function of time and temperature. METHODS: Eyes were obtained from minipigs and from human donors post mortem. Enucleated minipig eyes were stored for defined durations at three different temperatures (37 °C, 21 °C, and 4 °C). In order to assess the viability of the tissue, we measured ganglion cell activity (spiking) with multielectrode arrays. RESULTS: Minipig retinal ganglion cell function was severely compromised after 2 hours of ischemia at body temperature. After 4 hours, ganglion cells did not fire action potentials anymore. However, at 21 °C, ganglion cell activity was maintained under ischemic conditions for up to 12 hours, and for at least 50 hours at 4 °C. In postmortem human retina, we recorded ganglion cell activity in retinas received up to 27 hours after death. CONCLUSIONS: Our results demonstrate that hypothermia greatly increases survival of retinal ganglion cells exposed to ischemia. These results might be relevant for the future treatment of retinal ischemia.

High definition urethral pressure profilometry: Evaluating a novel microtip catheter.

INTRODUCTION: Urethral pressure profilometry (UPP) is used in the diagnosis of stress urinary incontinence (SUI). SUI is a significant medical, social, and economic problem, affecting about 12.5% of the population. A novel microtip catheter was developed for UPP featuring an inclination sensor and higher angular resolution compared to systems in clinical use today. Therewith, the location of each measured pressure sample can be determined and the spatial pressure distribution inside the urethra reconstructed. In order to assess the performance and plausibility of data from the microtip catheter, we compare it to data from a double balloon air charged system. MATERIALS AND METHODS: Both catheters are used on sedated female minipigs. Data from the microtip catheter are processed through a signal reconstruction algorithm, plotted and compared against data from the air-charged catheter. RESULTS: The microtip catheter delivers results in agreement with previous comparisons of microtip and air-charged systems. It additionally provides a new level of detail in the reconstructed UPPs which may lead to new insights into the sphincter mechanism of minipigs. CONCLUSIONS: The ability of air-charged catheters to measure pressure circumferentially is widely considered a main advantage over microtip catheters. However, directional pressure readings can provide additional information on angular fluctuations in the urethral pressure distribution. It is shown that the novel microtip catheter in combination with a signal reconstruction algorithm delivers plausible data. It offers the opportunity to evaluate urethral structures, especially the sphincter, in context of the correct location within the anatomical location of the pelvic floor. Neurourol. Urodynam. 35:888-894, 2016. © 2015 Wiley Periodicals, Inc.

Viability and MR detectability of iron labeled mesenchymal stem cells used for endoscopic injection into the porcine urethral sphincter.

Direct stem cell therapies for functionally impaired tissue require a sufficient number of cells in the target region and a method for verifying the fate of the cells in the subsequent time course. In vivo MRI of iron labeled mesenchymal stem cells has been suggested to comply with these requirements. The study was conducted to evaluate proliferation, migration, differentiation and adhesion effects as well as the obtained iron load of an iron labeling strategy for mesenchymal stem cells. After injection into the porcine urethral sphincter, the labeled cells were monitored for up to six months using MRI. Mesenchymal stem cells were labeled with ferucarbotran (60/100/200 µg/mL) and ferumoxide (200 µg/mL) for the analysis of migration and viability. Phantom MR measurements were made to evaluate effects of iron labeling. For short and long term studies, the iron labeled cells were injected into the porcine urethral sphincter and monitored by MRI. High resolution anatomical images of the porcine urethral sphincter were applied for detection of the iron particles with a turbo-spin-echo sequence and a gradient-echo sequence with multiple TE values. The MR images were then compared with histological staining. The analysis of cell function after iron labeling showed no effects on proliferation or differentiation of the cells. Although the adherence increases with higher iron dose, the ability to migrate decreases as a presumed effect of iron labeling. The iron labeled mesenchymal stem cells were detectable in vivo in MRI and histological staining even six months after injection. Labeling with iron particles and subsequent evaluation with highly resolved three dimensional data acquisition allows sensitive tracking of cells injected into the porcine urethral sphincter for several months without substantial biological effects on mesenchymal stem cells.

PDE3A mutations cause autosomal dominant hypertension with brachydactyly.

Cardiovascular disease is the most common cause of death worldwide, and hypertension is the major risk factor. Mendelian hypertension elucidates mechanisms of blood pressure regulation. Here we report six missense mutations in PDE3A (encoding phosphodiesterase 3A) in six unrelated families with mendelian hypertension and brachydactyly type E (HTNB). The syndrome features brachydactyly type E (BDE), severe salt-independent but age-dependent hypertension, an increased fibroblast growth rate, neurovascular contact at the rostral-ventrolateral medulla, altered baroreflex blood pressure regulation and death from stroke before age 50 years when untreated. In vitro analyses of mesenchymal stem cell-derived vascular smooth muscle cells (VSMCs) and chondrocytes provided insights into molecular pathogenesis. The mutations increased protein kinase A-mediated PDE3A phosphorylation and resulted in gain of function, with increased cAMP-hydrolytic activity and enhanced cell proliferation. Levels of phosphorylated VASP were diminished, and PTHrP levels were dysregulated. We suggest that the identified PDE3A mutations cause the syndrome. VSMC-expressed PDE3A deserves scrutiny as a therapeutic target for the treatment of hypertension.

Regeneration of degenerated urinary sphincter muscles: improved stem cell-based therapies and novel imaging technologies.

Stress urinary incontinence (SUI) is a largely ousted but significant medical, social, and economic problem. Surveys suggest that nowadays approximately 10% of the male and 15% of the female population suffer from urinary incontinence at some stage in their lifetime. In women, two major etiologies contribute to SUI: degeneration of the urethral sphincter muscle controlling the closing mechanism of the bladder outflow and changes in lower pelvic organ position associated with degeneration of connective tissue or with mechanical stress, including obesity and load and tissue injury during pregnancy and delivery. In males, the reduction of the sphincter muscle function is sometimes due to surgical interventions as a consequence of prostate cancer treatment, benign prostate hyperplasia, or of neuropathical origin. Accordingly, for women and men different therapies were developed. In some cases, SUI can be treated by physical exercise, electrophysiological stimulation, and pharmacological interventions. If this fails to improve the situation, surgical interventions are required. In standard procedures, endoprostheses for mechanical support of the weakened tissue or mechanical valves for a bladder outflow control are implanted. In 20% of cases treated, repeat procedures are required as implants yield all sorts of side effects in time. Based on preclinical studies, the application of an advanced therapy medicinal product (ATMP) such as implantation of autologous cells may be a curative and long-lasting therapy for SUI. Cellular therapy could also be an option for men suffering from incontinence caused by injury of the nerves controlling the muscular sphincter system. Here we briefly report on human progenitor cells, especially on mesenchymal stromal cells (MSCs), their expansion and differentiation to smooth muscle or striated muscle cells in vitro, labeling of cells for in vivo imaging, concepts of improved, precise, yet gentle application of cells in muscle tissue, and monitoring of injected cells in situ.

Tissue engineering in urothelium regeneration.

The development of therapeutic treatments to regenerate urothelium, manufacture tissue equivalents or neourethras for in-vivo application is a significant challenge in the field of tissue engineering. Many studies have focused on urethral defects that, in most cases, inadequately address current therapies. This article reviews the primary tissue engineering strategies aimed at the clinical requirements for urothelium regeneration while concentrating on promising investigations in the use of grafts, cellular preparations, as well as seeded or unseeded natural and synthetic materials. Despite significant progress being made in the development of scaffolds and matrices, buccal mucosa transplants have not been replaced. Recently, graft tissues appear to have an advantage over the use of matrices. These therapies depend on cell isolation and propagation in vitro that require, not only substantial laboratory resources, but also subsequent surgical implant procedures. The choice of the correct cell source is crucial when determining an in-vivo application because of the risks of tissue changes and abnormalities that may result in donor site morbidity. Addressing an appropriately-designed animal model and relevant regulatory issues is of fundamental importance for the principal investigators when a therapy using cellular components has been developed for clinical use.

Labelling and tracking of human mesenchymal stromal cells in preclinical studies and large animal models of degenerative diseases.

Success of stem cell therapies were reported in different medical disciplines, including haematology, rheumatology, orthopaedic surgery, traumatology, and others. Currently, more than 4000 clinical trials using stem cells have been completed or are underway, among which 378 investigated or are at present investigating mesenchymal stromal cells (MSCs). The majority of clinical trials using stem- or progenitor- cells, including hematopoietic stem cells and MSCs, target the immune system. However, therapies based on MSCs are increasingly implemented to treat symptoms in which failure of the resident stem cells in situ, or malfunction of tissues or structures are not associated with immune cells or inflammation, but instead are associated with mechanical or metabolic stress, ageing, developmental or acquired malformations, and other causes. To proceed further in the development of stem cell therapies as a safe and effective treatment for surgical and other medical specialities, the behaviour of MSCs implanted in preclinical models and their impact on the site of application need to be explored in detail. Depending on the pre-clinical model employed, tracking of labelled stem cells in live animals makes an enormous difference for exploration of the mechanisms and kinetics involved in MSC-mediated tissue regeneration. Here we review (pre-)clinically applicable key methods to label human MSCs for short and long-term observations in small and large animal models.

Towards a Treatment of Stress Urinary Incontinence: Application of Mesenchymal Stromal Cells for Regeneration of the Sphincter Muscle.

Stress urinary incontinence is a significant social, medical, and economic problem. It is caused, at least in part, by degeneration of the sphincter muscle controlling the tightness of the urinary bladder. This muscular degeneration is characterized by a loss of muscle cells and a surplus of a fibrous connective tissue. In Western countries approximately 15% of all females and 10% of males are affected. The incidence is significantly higher among senior citizens, and more than 25% of the elderly suffer from incontinence. When other therapies, such as physical exercise, pharmacological intervention, or electrophysiological stimulation of the sphincter fail to improve the patient's conditions, a cell-based therapy may improve the function of the sphincter muscle. Here, we briefly summarize current knowledge on stem cells suitable for therapy of urinary incontinence: mesenchymal stromal cells, urine-derived stem cells, and muscle-derived satellite cells. In addition, we report on ways to improve techniques for surgical navigation, injection of cells in the sphincter muscle, sensors for evaluation of post-treatment therapeutic outcome, and perspectives derived from recent pre-clinical studies.

Cell-based therapy for the deficient urinary sphincter.

When sterile culture techniques of mammalian cells first became state of the art, there was tremendous anticipation that such cells could be eventually applied for therapeutic purposes. The discovery of adult human stem or progenitor cells further motivated scientists to pursue research in cell-based therapies. Although evidence from animal studies suggests that application of cells yields measurable benefits, in urology and many other disciplines, progenitor-cell-based therapies are not yet routinely clinically available. Stress urinary incontinence (SUI) is a condition affecting a large number of patients. The etiology of SUI includes, but is not limited to, degeneration of the urinary sphincter muscle tissue and loss of innervation, as well as anatomical and biomechanical causes. Therefore, different regimens were developed to treat SUI. However, at present, a curative functional treatment is not at hand. A progenitor-cell-based therapy that can tackle the etiology of incontinence, rather than the consequences, is a promising strategy. Therefore, several research teams have intensified their efforts to develop such a therapy for incontinence. Here, we introduce candidate stem and progenitor cells suitable for SUI treatment, show how the functional homogeneity and state of maturity of differentiated cells crucial for proper tissue integration can be assessed electrophysiologically prior to their clinical application, and discuss the trophic potential of adult mesenchymal stromal (or stem) cells in regeneration of neuronal function.

Stem cell therapy for voiding and erectile dysfunction.

Voiding dysfunction comprises a variety of disorders, including stress urinary incontinence and overactive bladder, and affects millions of men and women worldwide. Erectile dysfunction (ED) also decreases quality of life for millions of men, as well as for their partners. Advanced age and diabetes are common comorbidities that can exacerbate and negatively impact upon the development of these disorders. Therapies that target the pathophysiology of these conditions to halt progression are not currently available. However, stem cell therapy could fill this therapeutic void. Stem cells can reduce inflammation, prevent fibrosis, promote angiogenesis, recruit endogenous progenitor cells, and differentiate to replace damaged cells. Adult multipotent stem cell therapy, in particular, has shown promise in case reports and preclinical animal studies. Stem cells also have a role in urological tissue engineering for ex vivo construction of bladder wall and urethral tissue (using a patient's own cells) prior to transplantation. More recent studies have focused on bioactive factor secretion and homing of stem cells. In the future, clinicians are likely to utilize allogeneic stem cell sources, intravenous systemic delivery, and ex vivo cell enhancement to treat voiding dysfunction and ED.

From tissue engineering to regenerative medicine in urology--the potential and the pitfalls.

Tissue engineering is a promising technique for the development of biological substitutes that can restore, maintain, or improve tissue function. The creation of human tissue-engineered products, generated of autologous somatic cells or adult stem cells with or without seeding of biocompatible matrices is a vision to resolve the lack of tissues and organs for transplantation and to offer new options for reconstructive surgery. Tissue engineering in urology aims at the reconstruction of the urinary tract by creating anatomically and functionally equal tissue. It is a rapidly evolving field in basic research and the transfer into the clinic has yet to be realized. Necessary steps from bench to bed are the proof of principle in animal models and the proof of concept in clinical trials following good manufacturing practice and ethical and legal requirements for human tissue-engineered products. Up to now, obstacles still occur in the neovascularization of implants and ingrowth of nerves in vivo. Moreover the harvesting of mesenchymal stem cells out of bone marrow as well as the explant of urothelial cells yet demands rather invasive surgery to achieve a successful outcome. Thus, other cell sources and harvesting techniques like placenta and adipose tissue for mesenchymal stem cells and bladder irrigation for urothelial cells require closer investigation.

Tumour stromal cells derived from paediatric malignancies display MSC-like properties and impair NK cell cytotoxicity.

BACKGROUND: Tumour growth and metastatic infiltration are favoured by several components of the tumour microenvironment. Bone marrow-derived multipotent mesenchymal stromal cells (MSC) are known to contribute to the tumour stroma. When isolated from healthy bone marrow, MSC exert potent antiproliferative effects on immune effector cells. Due to phenotypic and morphological similarities of MSC and tumour stromal cells (TStrC), we speculated that immunotherapeutic approaches may be hampered if TStrC may still exhibit immunomodulatory properties of MSC. METHODS: In order to compare immunomodulatory properties of MSC and tumour stromal cells (TStrC), we established and analyzed TStrC cultures from eleven paediatric tumours and MSC preparations from bone marrow aspirates. Immunophenotyping, proliferation assays and NK cell cytotoxicity assays were employed to address the issue. RESULTS: While TStrC differed from MSC in terms of plasticity, they shared surface expression of CD105, CD73 and other markers used for MSC characterization. Furthermore, TStrC displayed a strong antiproliferative effect on peripheral blood mononuclear cells (PBMC) in coculture experiments similar to MSC. NK cell cytotoxicity was significantly impaired after co-culture with TStrC and expression of the activating NK cell receptors NKp44 and NKp46 was reduced. CONCLUSIONS: Our data show that TStrC and MSC share important phenotypic and functional characteristics. The inhibitory effect of TStrC on PBMC and especially on NK cells may facilitate the immune evasion of paediatric tumours.

Low physiologic oxygen tensions reduce proliferation and differentiation of human multipotent mesenchymal stromal cells.

BACKGROUND: Human multipotent mesenchymal stromal cells (MSC) can be isolated from various tissues including bone marrow. Here, MSC participate as bone lining cells in the formation of the hematopoietic stem cell niche. In this compartment, the oxygen tension is low and oxygen partial pressure is estimated to range from 1% to 7%. We analyzed the effect of low oxygen tensions on human MSC cultured with platelet-lysate supplemented media and assessed proliferation, morphology, chromosomal stability, immunophenotype and plasticity. RESULTS: After transferring MSC from atmospheric oxygen levels of 21% to 1%, HIF-1alpha expression was induced, indicating efficient oxygen reduction. Simultaneously, MSC exhibited a significantly different morphology with shorter extensions and broader cell bodies. MSC did not proliferate as rapidly as under 21% oxygen and accumulated in G1 phase. The immunophenotype, however, was unaffected. Hypoxic stress as well as free oxygen radicals may affect chromosomal stability. However, no chromosomal abnormalities in human MSC under either culture condition were detected using high-resolution matrix-based comparative genomic hybridization. Reduced oxygen tension severely impaired adipogenic and osteogenic differentiation of human MSC. Elevation of oxygen from 1% to 3% restored osteogenic differentiation. CONCLUSION: Physiologic oxygen tension during in vitro culture of human MSC slows down cell cycle progression and differentiation. Under physiological conditions this may keep a proportion of MSC in a resting state. Further studies are needed to analyze these aspects of MSC in tissue regeneration.

Mesenchymal stromal cells: a novel treatment option for steroid-induced avascular osteonecrosis.

Mesenchymal stromal cells are multipotent cells capable of tissue repair and immune modulation. They are primarily found in bone marrow, but are also present in other tissues of mesenchymal origin, such as fatty tissue, muscle, tendons, etc. MSC can easily be obtained by bone marrow aspiration, showing a rapid expansion in vitro. New protocols enable cell culture without the use of animal-derived sera and artificial growth factors. Avascular necroses of the bone may have different causes. AVN in autoimmune and hematological diseases show a strong association with corticosteroid treatment, which is often unavoidable in severe cases. Until recently, core decompression of the affected osseous area was the standard approach. Because of their differentiation properties, easy accessibility and proliferative capacity, autologous MSCs could potentially complement AVN treatment by adding fresh "osteogenic cells" to the healing process.