Transplanted human neural stem cells rescue phenotypes in zQ175 Huntington's disease mice and innervate the striatum.

Huntington's disease (HD), a genetic neurodegenerative disorder, primarily affects the striatum and cortex with progressive loss of medium-sized spiny neurons (MSNs) and pyramidal neurons, disrupting cortico-striatal circuitry. A promising regenerative therapeutic strategy of transplanting human neural stem cells (hNSCs) is challenged by the need for long-term functional integration. We previously described that, with short-term hNSC transplantation into the striatum of HD R6/2 mice, human cells differentiated into electrophysiologically active immature neurons, improving behavior and biochemical deficits. Here, we show that long-term (8 months) implantation of hNSCs into the striatum of HD zQ175 mice ameliorates behavioral deficits, increases brain-derived neurotrophic factor (BDNF) levels, and reduces mutant huntingtin (mHTT) accumulation. Patch clamp recordings, immunohistochemistry, single-nucleus RNA sequencing (RNA-seq), and electron microscopy demonstrate that hNSCs differentiate into diverse neuronal populations, including MSN- and interneuron-like cells, and form connections. Single-nucleus RNA-seq analysis also shows restoration of several mHTT-mediated transcriptional changes of endogenous striatal HD mouse cells. Remarkably, engrafted cells receive synaptic inputs, innervate host neurons, and improve membrane and synaptic properties. Overall, the findings support hNSC transplantation for further evaluation and clinical development for HD.

Challenges of translating a cell therapy to GMP.

Over the last decade, cell and gene therapies have contributed remarkably to the array of novel therapies combating diseases that did not have any hope for an effective treatment or, let alone, a cure. This remarkable achievement was underlined by the marketing approval of CAR T cell therapies in 2017 in the United States, followed by many other countries, world-wide. Since then, thousands of patients have benefited from this autologous, gene modified cell therapy (Abou-El-Enein et al., 2021). Rare diseases, particularly innate neurological diseases such as Huntington's disease have also been a target for cell therapies. The notion of being able to augment or replace the function of diseased neurons with progenitor cells or neurons derived from human stem cells has been researched for the last 10 years and is finally reaching the stage of clinical translation (Holley et al., 2018; Reidling et al., 2018). With these cellular and gene therapies reaching clinical applicability, it is important to bring them to patients in a safe, efficacious and reliable way, and for this purpose, Good Manufacturing Practice (GMP) needs to be applied to the manufacturing of such novel and often life-saving therapies. In the first decade of the 21st century, gene therapies, particularly in vivo adenoviral vector gene therapy (Wilson, 2009) and hematopoietic stem cell gene therapies (Hacein-Bey-Abina et al., 2008) were associated with adverse events that were highly publicized and gave the field a bad reputation in the public eye. The last two decades, however, due to the meticulous work of dedicated researchers, and excellent progress in GMP manufacturing, cell and gene therapies have become safe and efficacious and have propelled the field to the forefront of the most promising novel therapies available for current unmet medical needs. This book chapter will discuss the historical perspective of cellular therapies and their development, will describe the currently available cell and gene therapies for different diseases and their GMP manufacturing methods and challenges, and will point out the future direction of these therapies and their envisioned manufacturing, as can be foreseen currently.

Target receptor identification and subsequent treatment of resected brain tumors with encapsulated and engineered allogeneic stem cells.

Cellular therapies offer a promising therapeutic strategy for the highly malignant brain tumor, glioblastoma (GBM). However, their clinical translation is limited by the lack of effective target identification and stringent testing in pre-clinical models that replicate standard treatment in GBM patients. In this study, we show the detection of cell surface death receptor (DR) target on CD146-enriched circulating tumor cells (CTC) captured from the blood of mice bearing GBM and patients diagnosed with GBM. Next, we developed allogeneic "off-the-shelf" clinical-grade bifunctional mesenchymal stem cells (MSCBif) expressing DR-targeted ligand and a safety kill switch. We show that biodegradable hydrogel encapsulated MSCBif (EnMSCBif) has a profound therapeutic efficacy in mice bearing patient-derived invasive, primary and recurrent GBM tumors following surgical resection. Activation of the kill switch enhances the efficacy of MSCBif and results in their elimination post-tumor treatment which can be tracked by positron emission tomography (PET) imaging. This study establishes a foundation towards a clinical trial of EnMSCBif in primary and recurrent GBM patients.

Scalable Manufacturing of CAR T cells for Cancer Immunotherapy.

As of April 2021, there are five commercially available chimeric antigen receptor (CAR) T cell therapies for hematological malignancies. With the current transition of CAR T cell manufacturing from academia to industry, there is a shift toward Good Manufacturing Practice (GMP)-compliant closed and automated systems to ensure reproducibility and to meet the increased demand for cancer patients. In this review we describe current CAR T cells clinical manufacturing models and discuss emerging technological advances that embrace scaling and production optimization. We summarize measures being used to shorten CAR T-cell manufacturing times and highlight regulatory challenges to scaling production for clinical use. STATEMENT OF SIGNIFICANCE ∣: As the demand for CAR T cell cancer therapy increases, several closed and automated production platforms are being deployed, and others are in development.This review provides a critical appraisal of these technologies that can be leveraged to scale and optimize the production of next generation CAR T cells.

Epileptic aphasia - A critical appraisal.

INTRODUCTION: Aphasic and other language disturbances occur in patients with epilepsy during and after epileptic seizures. Moreover, the interictal language profile in these patients is heterogeneous, varying from normal language profile to impairment in different language functions. The aim of this paper was to critically review the terms and concepts of ictal language alterations. MATERIAL AND METHOD: For this review we performed an extensive literature search on the term "epileptic aphasia" and analyzed the semiology and terminology indicating language-associated seizure symptoms. In addition, we give an overview on EEG, etiology, and brain imaging findings and ictal language disorders. RESULTS: In the literature, a plethora of terms indicates language-associated seizure symptoms. Simultaneous Video-EEG monitoring represents the gold standard to correctly classify ictal versus postictal language disturbances and to differentiate aphasic symptoms from speech automatisms. Different rhythmic and periodic EEG patterns associated with ictal language disturbances are recognized. Cerebral magnetic resonance imaging (cMRI) is essential in the diagnosis of seizures and epilepsy. Brain tumors and acute or remote cerebrovascular lesions are the most frequently reported structural etiologies underlying ictal language alterations. However, it has to be recognized that brain imaging may show alterations being the consequence of seizures itself rather than its cause. Functional brain imaging might be informative in patients with inconclusive EEG and MRI findings. Overall, seizure-associated aphasia is reported to have good lateralizing significance. CONCLUSION: Various language disturbances are caused by different types of seizures, epilepsies and underlying etiologies. In the clinical context, simultaneous Video-EEG monitoring facilitates precise classification of ictal versus postictal language alterations and differentiation of aphasic symptoms from speech automatisms.

Repetitive miniature spikes - An underreported EEG pattern.

OBJECTIVES: Low-voltage repetitive spikes are mainly described with invasive recordings and considered highly suggestive for focal cortical dysplasia (FCD). This EEG pattern has received less attention in routine scalp EEG. METHODS: Prospective collection of EEGs with low-voltage (<50 µV) repetitive spikes (repetitive miniature spikes - RMS) between July 1982 and July 2017 at the EEG laboratory of the Medical University of Innsbruck. We analyzed patterns of RMS on routine scalp EEG recordings and examined the relationship to clinical and brain imaging data. RESULTS: Overall, RMS were seen in 38 patients representing zero to four observations out of 5000 records per year. RMS occurred rhythmically in 14, periodically in 17 and irregularly in seven patients. The EEG pattern appeared with a frontal and central predominance. All but five patients had epilepsies; eleven patients had non-convulsive status epilepticus. Cerebral magnetic resonance imaging (cMRI) detected malformations of cortical development in eleven patients, including six patients with focal cortical dysplasias. CONCLUSIONS: RMS are rare EEG patterns indicating focal epilepsy. Their observation on routine scalp EEGs should prompt further clinic-radiologic investigation. SIGNIFICANCE: RMS resemble a clearly recognizable pattern in routine EEG, which is highly associated with focal epilepsy. The term is descriptive and can be added to the red flags, which can be found on routine EEG indicating underlying structural brain pathology, often in form of focal cortical dysplasia.

Primed mesenchymal stem cells package exosomes with metabolites associated with immunomodulation.

Mesenchymal stem cell (MSC) based therapies are currently being evaluated as a putative therapeutic in numerous human clinical trials. Recent reports have established that exosomes mediate much of the therapeutic properties of MSCs. Exosomes are nanovesicles which mediate intercellular communication, transmitting signals between cells which regulate a diverse range of biological processes. MSC-derived exosomes are packaged with numerous types of proteins and RNAs, however, their metabolomic and lipidomic profiles to date have not been well characterized. We previously reported that MSCs, in response to priming culture conditions that mimic the in vivo microenvironmental niche, substantially modulate cellular signaling and significantly increase the secretion of exosomes. Here we report that MSCs exposed to such priming conditions undergo glycolytic reprogramming, which homogenizes MSCs' metabolomic profile. In addition, we establish that exosomes derive from primed MSCs are packaged with numerous metabolites that have been directly associated with immunomodulation, including M2 macrophage polarization and regulatory T lymphocyte induction.

Exosomes Derived from Human Primed Mesenchymal Stem Cells Induce Mitosis and Potentiate Growth Factor Secretion.

Mesenchymal stem cells (MSCs) facilitate functional recovery in numerous animal models of inflammatory and ischemic tissue-related diseases with a growing body of research suggesting that exosomes mediate many of these therapeutic effects. It remains unclear, however, which types of proteins are packaged into exosomes compared with the cells from which they are derived. In this study, using comprehensive proteomic analysis, we demonstrated that human primed MSCs secrete exosomes (pMEX) that are packaged with markedly higher fractions of specific protein subclasses compared with their cells of origin, indicating regulation of their contents. Notably, we found that pMEX are also packaged with substantially elevated levels of extracellular-associated proteins. Fibronectin was the most abundant protein detected, and data established that fibronectin mediates the mitogenic properties of pMEX. In addition, treatment of SHSY5Y cells with pMEX induced the secretion of growth factors known to possess mitogenic and neurotrophic properties. Taken together, our comprehensive analysis indicates that pMEX are packaged with specific protein subtypes, which may provide a molecular basis for their distinct functional properties.

Mesenchymal stem/stromal cells genetically engineered to produce vascular endothelial growth factor for revascularization in wound healing and ischemic conditions.

Mesenchymal stem/stromal cells (MSCs) may be able to improve ischemic conditions as they can actively seek out areas of low oxygen and secrete proangiogenic factors. In more severe trauma and chronic cases, however, cells alone may not be enough. Therefore, we have combined the stem cell and angiogenic factor approaches to make a more potent therapy. We developed an engineered stem cell therapy product designed to treat critical limb ischemia that could also be used in trauma-induced scarring and fibrosis where additional collateral blood flow is needed following damage to and blockage of the primary vessels. We used MSCs from normal human donor marrow and engineered them to produce high levels of the angiogenic factor vascular endothelial growth factor (VEGF). The MSC/VEGF product has been successfully developed and characterized using good manufacturing practice (GMP)-compliant methods, and we have completed experiments showing that MSC/VEGF significantly increased blood flow in the ischemic limb of immune deficient mice, compared to the saline controls in each study. We also performed safety studies demonstrating that the injected product does not cause harm and that the cells remain around the injection site for more than 1 month after hypoxic preconditioning. An on-demand formulation system for delivery of the product to clinical sites that lack cell processing facilities is in development.

Concise Review: A Comprehensive Analysis of Reported Adverse Events in Patients Receiving Unproven Stem Cell-Based Interventions.

The promise of stem cell (SC) therapies to restore functions of damaged tissues and organs brings enormous hope to patients, their families, loved ones, and caregivers. However, limits may exist for which indications SC therapies might be useful, efficacious, and safe. Applications of innovative therapies within regulatory boundaries and within the framework of controlled clinical trials are the norm in the scientific and medical community; such a system minimizes patient risk by setting a clear and acceptable safety and efficacy profile for new therapeutics before marketing authorization. This careful clinical validation approach often takes time, which patients suffering from terminal or debilitating diseases do not have. Not validated, unproven stem cell interventions (SCI) that promise a working treatment or cure for severe diseases have therefore found their way into the patient community, and providers of such treatments often take advantage of the public's willingness to pay large amounts of money for the misguided hope of a reliable recovery from their illnesses. We conducted a review of scientific publications, clinical case reports, and mass media publications to assess the reported cases and safety incidents associated with unproven SCI. The review also analyzes the main factors that were identified as contributing to the emergence and global rise of the "stem cell tourism" phenomenon. Stem Cells Translational Medicine 2018;1-10.

SV-BR-1-GM, a Clinically Effective GM-CSF-Secreting Breast Cancer Cell Line, Expresses an Immune Signature and Directly Activates CD4+ T Lymphocytes.

Targeted cancer immunotherapy with irradiated, granulocyte-macrophage colony-stimulating factor (GM-CSF)-secreting, allogeneic cancer cell lines has been an effective approach to reduce tumor burden in several patients. It is generally assumed that to be effective, these cell lines need to express immunogenic antigens coexpressed in patient tumor cells, and antigen-presenting cells need to take up such antigens then present them to patient T cells. We have previously reported that, in a phase I pilot study (ClinicalTrials.gov NCT00095862), a subject with stage IV breast cancer experienced substantial regression of breast, lung, and brain lesions following inoculation with clinical formulations of SV-BR-1-GM, a GM-CSF-secreting breast tumor cell line. To identify diagnostic features permitting the prospective identification of patients likely to benefit from SV-BR-1-GM, we conducted a molecular analysis of the SV-BR-1-GM cell line and of patient-derived blood, as well as a tumor specimen. Compared to normal human breast cells, SV-BR-1-GM cells overexpress genes encoding tumor-associated antigens (TAAs) such as PRAME, a cancer/testis antigen. Curiously, despite its presumptive breast epithelial origin, the cell line expresses major histocompatibility complex (MHC) class II genes (HLA-DRA, HLA-DRB3, HLA-DMA, HLA-DMB), in addition to several other factors known to play immunostimulatory roles. These factors include MHC class I components (B2M, HLA-A, HLA-B), ADA (encoding adenosine deaminase), ADGRE5 (CD97), CD58 (LFA3), CD74 (encoding invariant chain and CLIP), CD83, CXCL8 (IL8), CXCL16, HLA-F, IL6, IL18, and KITLG. Moreover, both SV-BR-1-GM cells and the responding study subject carried an HLA-DRB3*02:02 allele, raising the question of whether SV-BR-1-GM cells can directly present endogenous antigens to T cells, thereby inducing a tumor-directed immune response. In support of this, SV-BR-1-GM cells (which also carry the HLA-DRB3*01:01 allele) treated with yellow fever virus (YFV) envelope (Env) 43-59 peptides reactivated YFV-DRB3*01:01-specific CD4+ T cells. Thus, the partial HLA allele match between SV-BR-1-GM and the clinical responder might have enabled patient T lymphocytes to directly recognize SV-BR-1-GM TAAs as presented on SV-BR-1-GM MHCs. Taken together, our findings are consistent with a potentially unique mechanism of action by which SV-BR-1-GM cells can act as APCs for previously primed CD4+ T cells.

Therapeutic effects of stem cells in rodent models of Huntington's disease: Review and electrophysiological findings.

The principal symptoms of Huntington's disease (HD), chorea, cognitive deficits, and psychiatric symptoms are associated with the massive loss of striatal and cortical projection neurons. As current drug therapies only partially alleviate symptoms, finding alternative treatments has become peremptory. Cell replacement using stem cells is a rapidly expanding field that offers such an alternative. In this review, we examine recent studies that use mesenchymal cells, as well as pluripotent, cell-derived products in animal models of HD. Additionally, we provide further electrophysiological characterization of a human neural stem cell line, ESI-017, which has already demonstrated disease-modifying properties in two mouse models of HD. Overall, the field of regenerative medicine represents a viable and promising avenue for the treatment of neurodegenerative disorders including HD.

Human Neural Stem Cell Transplantation Rescues Functional Deficits in R6/2 and Q140 Huntington's Disease Mice.

Huntington's disease (HD) is an inherited neurodegenerative disorder with no disease-modifying treatment. Expansion of the glutamine-encoding repeat in the Huntingtin (HTT) gene causes broad effects that are a challenge for single treatment strategies. Strategies based on human stem cells offer a promising option. We evaluated efficacy of transplanting a good manufacturing practice (GMP)-grade human embryonic stem cell-derived neural stem cell (hNSC) line into striatum of HD modeled mice. In HD fragment model R6/2 mice, transplants improve motor deficits, rescue synaptic alterations, and are contacted by nerve terminals from mouse cells. Furthermore, implanted hNSCs are electrophysiologically active. hNSCs also improved motor and late-stage cognitive impairment in a second HD model, Q140 knockin mice. Disease-modifying activity is suggested by the reduction of aberrant accumulation of mutant HTT protein and expression of brain-derived neurotrophic factor (BDNF) in both models. These findings hold promise for future development of stem cell-based therapies.

ACL double-bundle reconstruction with one tibial tunnel provides equal stability compared to two tibial tunnels.

PURPOSE: The purpose of this study was to investigate whether an anterior cruciate ligament (ACL) double-bundle reconstruction with one tibial tunnel displays the same in vitro stability as a conventional double-bundle reconstruction with two tibial tunnels when using the same tensioning protocol. METHODS: In 11 fresh-frozen cadaveric knees, ACL double-bundle reconstruction with one and two tibial tunnels was performed. The two grafts were tightened using 80 N in different flexion angles (anteromedial-bundle at 60° and posterolateral-bundle at 15°). Anterior tibial translation (134 N) and translation with combined rotatory and valgus loads (10 Nm valgus stress and 4 Nm internal tibial torque) were determined at 0°, 30°, 60° and 90° flexion. Measurements were taken in intact ACL, resected ACL, three-tunnel reconstruction and four-tunnel reconstruction. Additionally, the tension on the grafts was determined. Student's t test was performed for statistical analysis of the related samples. Significance was set at p < 0.017 according to Bonferroni correction. RESULTS: The two reconstructive techniques displayed no significant differences in comparison with the intact ACL in anterior tibial translation at 0°, 60° and 90° of flexion. The same results were obtained for the anterior tibial translation with a combined rotatory load at 60° and 90°. When directly comparing both reconstructive techniques, there were no significant differences for the anterior tibial translation and combined rotatory load at all flexion angles. The measured tension on grafts displayed similar load sharing between both bundles. Except at full extension, both grafts displayed a significantly different tension increase under anterior tibial translation for both techniques (p = 0.0086). CONCLUSIONS: Tightening both bundles in ACL double-bundle reconstruction with one or two tibial tunnels in different flexion angles achieved comparable restoration of stability, although there was different load sharing on the bundles. With regard to individualized ACL reconstruction, the double-bundle technique with one tibial tunnel offers a possibility to address small tibial insertion sites without compromising the advantages of a double-bundle procedure.

Advances in bone marrow stem cell therapy for retinal dysfunction.

The most common cause of untreatable vision loss is dysfunction of the retina. Conditions, such as age-related macular degeneration, diabetic retinopathy and glaucoma remain leading causes of untreatable blindness worldwide. Various stem cell approaches are being explored for treatment of retinal regeneration. The rationale for using bone marrow stem cells to treat retinal dysfunction is based on preclinical evidence showing that bone marrow stem cells can rescue degenerating and ischemic retina. These stem cells have primarily paracrine trophic effects although some cells can directly incorporate into damaged tissue. Since the paracrine trophic effects can have regenerative effects on multiple cells in the retina, the use of this cell therapy is not limited to a particular retinal condition. Autologous bone marrow-derived stem cells are being explored in early clinical trials as therapy for various retinal conditions. These bone marrow stem cells include mesenchymal stem cells, mononuclear cells and CD34+ cells. Autologous therapy requires no systemic immunosuppression or donor matching. Intravitreal delivery of CD34+ cells and mononuclear cells appears to be tolerated and is being explored since some of these cells can home into the damaged retina after intravitreal administration. The safety of intravitreal delivery of mesenchymal stem cells has not been well established. This review provides an update of the current evidence in support of the use of bone marrow stem cells as treatment for retinal dysfunction. The potential limitations and complications of using certain forms of bone marrow stem cells as therapy are discussed. Future directions of research include methods to optimize the therapeutic potential of these stem cells, non-cellular alternatives using extracellular vesicles, and in vivo high-resolution retinal imaging to detect cellular changes in the retina following cell therapy.

Putting a price tag on novel autologous cellular therapies.

Cell therapies, especially autologous therapies, pose significant challenges to researchers who wish to move from small, probably academic, methods of manufacture to full commercial scale. There is a dearth of reliable information about the costs of operation, and this makes it difficult to predict with confidence the investment needed to translate the innovations to the clinic, other than as small-scale, clinician-led prescriptions. Here, we provide an example of the results of a cost model that takes into account the fixed and variable costs of manufacture of one such therapy. We also highlight the different factors that influence the product final pricing strategy. Our findings illustrate the need for cooperative and collective action by the research community in pre-competitive research to generate the operational models that are much needed to increase confidence in process development for these advanced products.

Comprehensive Proteomic Analysis of Mesenchymal Stem Cell Exosomes Reveals Modulation of Angiogenesis via Nuclear Factor-KappaB Signaling.

Mesenchymal stem cells (MSC) are known to facilitate healing of ischemic tissue related diseases through proangiogenic secretory proteins. Recent studies further show that MSC derived exosomes function as paracrine effectors of angiogenesis, however, the identity of which components of the exosome proteome responsible for this effect remains elusive. To address this we used high-resolution isoelectric focusing coupled liquid chromatography tandem mass spectrometry, an unbiased high throughput proteomics approach to comprehensively characterize the proteinaceous contents of MSCs and MSC derived exosomes. We probed the proteome of MSCs and MSC derived exosomes from cells cultured under expansion conditions and under ischemic tissue simulated conditions to elucidate key angiogenic paracrine effectors present and potentially differentially expressed in these conditions. In total, 6,342 proteins were identified in MSCs and 1,927 proteins in MSC derived exosomes, representing to our knowledge the first time these proteomes have been probed comprehensively. Multilayered analyses identified several putative paracrine effectors of angiogenesis present in MSC exosomes and increased in expression in MSCs exposed to ischemic tissue-simulated conditions; these include platelet derived growth factor, epidermal growth factor, fibroblast growth factor, and most notably nuclear factor-kappaB (NFkB) signaling pathway proteins. NFkB signaling was identified as a key mediator of MSC exosome induced angiogenesis in endothelial cells by functional in vitro validation using a specific inhibitor. Collectively, the results of our proteomic analysis show that MSC derived exosomes contain a robust profile of angiogenic paracrine effectors, which have potential for the treatment of ischemic tissue-related diseases.

Efficient Generation of Induced Pluripotent Stem and Neural Progenitor Cells From Acutely Harvested Dura Mater Obtained During Ventriculoperitoneal Shunt Surgery.

BACKGROUND: The dura mater can be easily biopsied during most cranial neurosurgical operations. We describe a protocol that allows for robust generation of induced pluripotent stem cells (iPSCs) and neural progenitors from acutely harvested dura mater. OBJECTIVE: To generate iPSCs and neural progenitor cells from dura mater obtained during ventriculoperitoneal shunt surgery. METHODS: Dura was obtained during ventriculoperitoneal shunt surgery for normal pressure hydrocephalus from a 60-year-old patient with severe cognitive impairment. Fibroblasts were isolated from the dural matrix and transduced with nonintegrating Sendai virus for iPSC induction. A subset of successfully generated iPSC clones underwent immunocytochemical analysis, teratoma assay, karyotyping, and targeted neural differentiation. RESULTS: Eleven iPSC clones were obtained from the transduction of an estimated 600,000 dural fibroblasts after 3 passages. Three clones underwent immunocytochemical analysis and were shown to express the transcription factors OCT-4, SOX2, and the embryonic cell markers SSEA-4, TRA-1-60, and Nanog. Two clones were tested for pluripotency and formed teratomas at the injection site in immunodeficient mice. Three clones underwent chromosomal analysis and were found to have a normal metaphase spread and karyotype. One clone underwent targeted neural differentiation and formed neural rosettes as well as TuJ1/SOX1-positive neural progenitor cells. CONCLUSIONS: IPSCs and neural progenitor cells can be efficiently derived from the dura of patients who need to undergo cranial neurosurgical operations. IPSCs were obtained with a nonintegrating virus and exhibited a normal karyotype, making them candidates for future autotransplantation after targeted differentiation to treat functional deficits.

Cause-specific mortality in adult epilepsy patients from Tyrol, Austria: hospital-based study.

Epilepsy is a devastating condition with a considerable increase in mortality compared to the general population. Few studies have focused on cause-specific mortality which we analyse in detail in over 4,000 well-characterized epilepsy patients. The cohort comprised of epilepsy patients ≥ 18, treated between 1970 and 2009 at the epilepsy clinic of Innsbruck Medical University, Austria, and living in the province of Tyrol, Austria. Epilepsy diagnosis was based on ILAE guidelines (1989); patients with brain tumor were excluded. Deceased patients and causes of death (ICD-codes) were obtained via record linkage to the national death registry. We computed age-, sex-, and period-adjusted standardized mortality rates (SMR) for 36 diagnoses subgroups in four major groups. Additional analyses were performed for an incidence cohort. Overall cohort: 4,295 patients, 60,649.1 person-years, 822 deaths, overall SMR 1.7 (95 % CI 1.6-1.9), highest elevated cause-specific SMR: congenital anomalies [7.1 (95 % CI 2.3-16.6)], suicide [4.2 (95 % CI 2.0-8.1)], alcohol dependence syndrome [3.9 (95 % CI 1.8-7.4)], malignant neoplasm of esophagus [3.1 (95 % CI 1.2-6.4)], pneumonia [2.7 (95 % CI 1.6-4.2)]. Incidence cohort: 1,299 patients, 14,215.4 person-years, 267 deaths, overall SMR 1.8 (95 % CI 1.6-2.1), highest elevated cause-specific SMR congenital anomalies [10.8 (95 % CI 1.3-39.3)], suicide [6.8 (95 % CI 1.4-19.8)], alcohol dependence syndrome (6.4 [95 % CI 1.8-16.5)], pneumonia [3.9 (95 % CI 1.8-7.4)], cerebrovascular disease at 3.5 (95 % CI 2.6-4.6). Mortality due to mental health problems, such as suicide or alcohol dependence syndrome, malignant neoplasms, and cerebrovascular diseases was highly increased in our study. In addition to aim for seizure freedom, we suggest improving general health promotion, including cessation of smoking, lowering of alcohol intake, and reduction of weight as well as early identification of psychiatric comorbidity in patients with epilepsy.

Safety and efficacy of a tCD25 preselective combination anti-HIV lentiviral vector in human hematopoietic stem and progenitor cells.

The successful suppression of human immunodeficiency virus (HIV) in the "Berlin Patient" has highlighted the ability of HIV-resistant hematopoietic stem cells to offer a potential functional cure for HIV-infected patients. HIV stem cell gene therapy can mimic this result by genetically modifying a patient's own cells with anti-HIV genes. Previous attempts of HIV gene therapy have been hampered by a low percentage of transplanted HIV-resistant cells which has led to minimal clinical efficacy. In our current study, we have evaluated the in vitro and in vivo safety and efficacy of a truncated/mutated form of human CD25 preselective anti-HIV lentiviral vector in human hematopoietic stem cells. This preselective vector allows us to purify vector-transduced cells prior to transplantation so an increased percentage of gene-modified cells can be delivered. Here, we demonstrate the safety of this strategy with successful engraftment and multilineage hematopoiesis of transduced cells in a humanized NOD-RAG1-/-IL-2rγ-/- knockout mouse model. Efficacy was also demonstrated with significant protection from HIV-1 infection including maintenance of human CD4+ cell levels and a decrease in HIV-1 plasma viremia. Collectively, these results establish the utility of this HIV stem cell gene therapy strategy and bring it closer to providing a functional cure for HIV-infected patients.