Immunogenicity, efficacy, safety, and mechanism of action of epitope vaccine (Lu AF20513) for Alzheimer's disease: prelude to a clinical trial.

The Alzheimer's disease (AD) process is understood to involve the accumulation of amyloid plaques and tau tangles in the brain. However, attempts at targeting the main culprits, neurotoxic Aß peptides, have thus far proven unsuccessful for improving cognitive function. Recent clinical trials with passively administrated anti-Aß antibodies failed to slow cognitive decline in mild to moderate AD patients, but suggest that an immunotherapeutic approach could be effective in patients with mild AD. Using an AD mouse model (Tg2576), we tested the immunogenicity (cellular and humoral immune responses) and efficacy (AD-like pathology) of clinical grade Lu AF20513 vaccine. We found that Lu AF20513 induces robust "non-self" T-cell responses and the production of anti-Aß antibodies that reduce AD-like pathology in the brains of Tg2576 mice without inducing microglial activation and enhancing astrocytosis or cerebral amyloid angiopathy. A single immunization with Lu AF20513 induced strong humoral immunity in mice with preexisting memory T-helper cells. In addition, Lu AF20513 induced strong humoral responses in guinea pigs and monkeys. These data support the translation of Lu AF20513 to the clinical setting with the aims of: (1) inducing therapeutically potent anti-Aß antibody responses in patients with mild AD, particularly if they have memory T-helper cells generated after immunizations with conventional tetanus toxoid vaccine, and (2) preventing pathological autoreactive T-cell responses.

Exosome removal as a therapeutic adjuvant in cancer.

Exosome secretion is a notable feature of malignancy owing to the roles of these nanoparticles in cancer growth, immune suppression, tumor angiogenesis and therapeutic resistance. Exosomes are 30-100 nm membrane vesicles released by many cells types during normal physiological processes. Tumors aberrantly secrete large quantities of exosomes that transport oncoproteins and immune suppressive molecules to support tumor growth and metastasis. The role of exosomes in intercellular signaling is exemplified by human epidermal growth factor receptor type 2 (HER2) over-expressing breast cancer, where exosomes with the HER2 oncoprotein stimulate tumor growth and interfere with the activity of the therapeutic antibody Herceptin®. Since numerous observations from experimental model systems point toward an important clinical impact of exosomes in cancer, several pharmacological strategies have been proposed for targeting their malignant activities. We also propose a novel device strategy involving extracorporeal hemofiltration of exosomes from the entire circulatory system using an affinity plasmapheresis platform known as the Aethlon ADAPT™ (adaptive dialysis-like affinity platform technology) system, which would overcome the risks of toxicity and drug interactions posed by pharmacological approaches. This technology allows affinity agents, including exosome-binding lectins and antibodies, to be immobilized in the outer-capillary space of plasma filtration membranes that integrate into existing kidney dialysis systems. Device therapies that evolve from this platform allow rapid extracorporeal capture and selective retention of target particles < 200 nm from the entire circulatory system. This strategy is supported by clinical experience in hepatitis C virus-infected patients using an ADAPT™ device, the Hemopurifier®, to reduce the systemic load of virions having similar sizes and glycosylated surfaces as cancer exosomes. This review discusses the possible therapeutic approaches for targeting immune suppressive exosomes in cancer patients, and the anticipated significance of these strategies for reversing immune dysfunction and improving responses to standard of care treatments.

IL-18 is required for self-reactive T cell expansion in NOD mice.

IL-18 has a well-established role in pro-inflammatory responses in the islets in type 1 diabetes. Here, we identify a distinctive role for IL-18 in expanding pathogenic T cells in the periphery of NOD mice. Well in advance of disease onset, the periphery of IL-18-deficient mice exhibits reduced T cell turnover, an increased prevalence of naïve and quiescent T cells, emergence of fewer effector T cells, and disease protection. Islet-reactive T cells fail to become activated in the lymphoid organs of mice lacking IL-18 and their rapid expansion is inhibited. IL-18 secretion by antigen presenting cells increases with advancing disease and is required for expression of its receptor on T cells. Our results demonstrate that induction of the IL-18 receptor reflects a critical stage of autoreactive T cell activation and expansion on the pathway toward effector T cell differentiation. This study therefore assigns a novel role to IL-18 for expanding the pool of islet-destructive T cells during pre-diabetes. This report highlights a new basic mechanism in type 1 diabetes pathogenesis and suggests that targeting the IL-18 pathway should be explored as a potential treatment strategy.

Autologous stromal vascular fraction cells: a tool for facilitating tolerance in rheumatic disease.

Since the days of Medawar, the goal of therapeutic tolerogenesis has been a "Holy Grail" for immunologists. While knowledge of cellular and molecular mechanisms of this process has been increasing at an exponential rate, clinical progress has been minimal. To provide a mechanistic background of tolerogenesis, we overview common processes in the naturally occurring examples of: pregnancy, cancer, oral tolerance and anterior chamber associated immune deviation. The case is made that an easily accessible byproduct of plastic surgery, the adipose stromal vascular fraction, contains elements directly capable of promoting tolerogenesis such as T regulatory cells and inhibitory macrophages. The high content of mesenchymal and hematopoietic stem cells from this source provides the possibility of trophic/regenerative potential, which would augment tolerogenic processes by decreasing ongoing inflammation. We discuss the application of this autologous cell source in the context of rheumatoid arthritis, concluding with some practical examples of its applications.

Mesenchymal stem cells as anti-inflammatories: implications for treatment of Duchenne muscular dystrophy.

Duchenne muscular dystrophy (DMD) is a lethal X-linked musculodegenerative condition consisting of an underlying genetic defect whose manifestation is augmented by inflammatory mechanisms. Previous treatment approaches using gene replacement, exon-skipping or allogeneic cell therapy have been relatively unsuccessful. The only intervention to mediate improvement in survival, albeit minor, is glucocorticoid treatment. Given this modality appears to function via suppression of underlying inflammation; we focus this review on the inflammatory response as a target for mesenchymal stem cell (MSC) therapy. In contrast to other cell based therapies attempted in DMD, MSC have the advantages of (a) ability to fuse with and genetically complement dystrophic muscle; (b) possess anti-inflammatory activities; and (c) produce trophic factors that may augment activity of endogenous repair cells. We conclude by describing one practical scenario of stem cell therapy for DMD.

Allogeneic endometrial regenerative cells: an "Off the shelf solution" for critical limb ischemia?

Critical limb ischemia (CLI) is an advanced form of peripheral artery disease which is responsible for approximately 100,000 amputations per year in the US. Trials to date have reported clinical improvement and reduced need for amputation in CLI patients receiving autologous bone marrow or mobilized peripheral blood stem cells for stimulation of angiogenesis. While such treatments are currently entering Phase III trials, practical and scientific pitfalls will limit widespread implementation if efficacy is proven. Hurdles to be overcome include: a) reduced angiogenic potential of autologous cells in aged patients with cardiovascular risk factors; b) invasiveness/adverse effects of bone marrow extraction and G-CSF mobilization, respectively; and c) need for on-site cellular manipulation. The Endometrial Regenerative Cell (ERC) is a mesenchymal-like stem cell derived from the menstrual blood that is believed to be associated with endometrial angiogenesis. We discuss the possibility of using allogeneic ERCs as an "off the shelf" treatment for CLI based on the following properties: a) High levels of growth factors and matrix metalloprotease production; b) Ability to inhibits inflammatory responses and lack of immunogenicity; and c) Expandability to great quantities without loss of differentiation ability or karyotypic abnormalities.

Immune effects of mesenchymal stem cells: implications for Charcot-Marie-Tooth disease.

Mesenchymal stem cell (MSC) therapy is the most clinically advanced form of cell therapy, second to hematopoietic stem cell transplants. To date, MSC have been used for immune modulation in conditions such as Graft Versus Host Disease (GVHD) and Crohn's Disease, for which Phase III clinical trials are currently in progress. Here, we review the immunological properties of MSC and make a case for their use in treatment of Charcot-Marie-Tooth disease type 1 (CMT1), a group of inherited peripheral neuropathies. CMT1 is characterized by demyelination and aberrant immune activation making this condition an ideal target for exploration of MSC therapy, given the ability of these cells to promote sheath regeneration as well as suppress inflammation. Studies supporting this hypothesis will be presented and placed into the context of other cell-based approaches that are theoretically feasible. Given that MSCs selectively home to areas of inflammation, as well as exert effects in an allogeneic manner, the possibility of an "off the shelf" therapy for CMT1 will be discussed.

Cord blood in regenerative medicine: do we need immune suppression?

Cord blood is currently used as an alternative to bone marrow as a source of stem cells for hematopoietic reconstitution after ablation. It is also under intense preclinical investigation for a variety of indications ranging from stroke, to limb ischemia, to myocardial regeneration. A major drawback in the current use of cord blood is that substantial morbidity and mortality are associated with pre-transplant ablation of the recipient hematopoietic system. Here we raise the possibility that due to unique immunological properties of both the stem cell and non-stem cell components of cord blood, it may be possible to utilize allogeneic cells for regenerative applications without needing to fully compromise the recipient immune system. Issues raised will include: graft versus host potential, the immunogenicity of the cord blood graft, and the parallels between cord blood transplantation and fetal to maternal trafficking. The previous use of unmatched cord blood in absence of any immune ablation, as well as potential steps for widespread clinical implementation of allogeneic cord blood grafts will also be discussed.

Therapeutic use of Aldara in chronic myeloid leukemia.

The potent clinical responses seen in patients with chronic myeloid leukemia (CML) after administration of donor-specific lymphocytes, as well as the correlation between the presence of antigen specific T cells and prolonged remission in these patients, suggests a role for the immunological control of CML. Here we propose Aldara, a clinically used formulation of imiquimod, as an agent for augmenting immune responses to CML antigens. Our proposition is based upon 3 tenets: 1) Endogenous dendritic cells (DC) of CML patients, which are known to be derived from the malignant clone, express and present various leukemic antigens; 2) CML-antigen reactive T cell clones exist in the patient but in many situations are ineffectively stimulated to cause significant hematological responses; and 3) Antigen presentation by mature, activated DC, which endogenously express CML-antigens may endow the pre-existing ineffective T cell responses with ability to control CML progression. The practical use of Aldara as a localized activator of DC in the context of present day leukemic therapeutics, as well as various properties of this unique immune modulator will be discussed.

T cell homeostasis in tolerance and immunity.

The size of the peripheral T cell pool is remarkably stable throughout life, reflecting precise regulation of cellular survival, proliferation, and apoptosis. Homeostatic proliferation refers to the process by which T cells spontaneously proliferate in a lymphopenic host. The critical signals driving this expansion are "space," contact with self-major histocompatibility complex (MHC)/peptide complexes, and cytokine stimulation. A number of studies have delineated an association between T cell lymphopenia, compensatory homeostatic expansion, and the development of diverse autoimmune syndromes. In the nonobese diabetic mouse model of type 1 diabetes, lymphopenia-induced homeostatic expansion fuels the generation of islet-specific T cells. Excess interleukin-21 facilitates T cell cycling but limited survival, resulting in recurrent stimulation of T cells specific for self-peptide/MHC complexes. Indeed, data from several experimental models of autoimmunity indicate that a full T cell compartment restrains homeostatic expansion of self-reactive cells that could otherwise dominate the repertoire. This review describes the mechanisms that govern T cell homeostatic expansion and outlines the evidence that lymphopenia presents a risk for development of autoimmune disease.

A dual vaccine against influenza & Alzheimer's disease failed to enhance anti-ß-amyloid antibody responses in mice with pre-existing virus specific memory.

Novel dual vaccine, WSN-Aß(1-10), based on the recombinant influenza virus, expressing immunodominant B-cell epitope of ß-amyloid, simultaneously induced therapeutically potent anti-Aß and anti-influenza antibodies. In this study we showed that boosting of WSN-WT primed mice with WSN-Aß(1-10) enhances anti-viral, but fails to induce anti-Aß antibody responses. This inhibition is associated with expression of Aß(1-10) within the context of an inactivated influenza virus vaccine. These results demonstrate that the use of an inactivated influenza virus as a carrier for AD vaccine may not be applicable due to the possible inhibition of anti-Aß antibody response in individuals previously vaccinated or infected with influenza.

Reduction of tumorigenicity by placental extracts.

The influence of adult stem cells on tumor growth is paradoxical. On one hand, angiogenic factors secreted by stem cells are known to be essential for tumor vascularization. On the other hand, stem cell-derived factors can reportedly induce tumor differentiation or direct death of tumor cells. Both the placenta and umbilical cord are rich sources of stem cells with immune modulatory and tissue-healing properties; however, the effects of placental components on cancer cells have not been fully defined. Here we demonstrate that extracts of placental lysates reduce the malignancy of a variety of human tumor cell lines in a species-unrestricted manner. Using a standard model of leukemia cell differentiation, we demonstrated that addition of placental extracts to tumor cells, or co-culture of tumor cells with the CD34(+) cells from umbilical cord blood, induced tumor cell differentiation. Inhibition of tumor growth and metastasis in vivo was also observed following administration of placental extracts. These data support the concept of non-toxic biological therapy of cancer using stem cell derivatives, possibly through the induction of tumor cell differentiation.

Autologous stromal vascular fraction therapy for rheumatoid arthritis: rationale and clinical safety.

Advancements in rheumatoid arthritis (RA) treatment protocols and introduction of targeted biological therapies have markedly improved patient outcomes, despite this, up to 50% of patients still fail to achieve a significant clinical response. In veterinary medicine, stem cell therapy in the form of autologous stromal vascular fraction (SVF) is an accepted therapeutic modality for degenerative conditions with 80% improvement and no serious treatment associated adverse events reported. Clinical translation of SVF therapy relies on confirmation of veterinary findings in targeted patient populations. Here we describe the rationale and preclinical data supporting the use of autologous SVF in treatment of RA, as well as provide 1, 3, 6, and 13 month safety outcomes in 13 RA patients treated with this approach.

Differential contributions of APC subsets to T cell activation in nonobese diabetic mice.

Despite the pivotal role of dendritic cells (DC) in shaping immunity, little is known about their functionality in type 1 diabetes. Moreover, due to the paucity of DC in vivo, functional studies have relied largely upon in vitro-expanded cells to elucidate type 1 diabetes-associated functional abnormalities. In this study, we provide a comprehensive analysis of the functional capabilities of in vivo-derived DC subsets from NOD mice by comparing DC to other NOD APC types and to DC from autoimmune-resistant strains. NOD DC closely resemble those from nonautoimmune strains with respect to costimulation and cytokine production. The exception is the CD8alpha(+)CD11b(-)DC subset which is numerically reduced in NOD spleens, but not in the pancreatic lymph nodes, while DC from both tissues produce little IL-12 in this strain. This defect results in unusual deferral toward macrophage-derived IL-12 in NOD mice; NOD macrophages produce aberrantly high IL-12 levels that can overcompensate for the DC defect in Th1 polarization. APC subset use for autoantigen presentation also differs in NOD mice. NOD B cells overshadow DC at activating islet-reactive T cells, whereas DC and B cells in NOD-resistant mice are functionally comparable. Differential involvement of APC subsets in T cell activation and tolerance induction may prove to be a crucial factor in the selection and expansion of autoreactive T cells.

Reduced IFN-alpha secretion by blood dendritic cells in human diabetes.

Characterization of dendritic cells (DC) in human diabetes has been restricted to monocyte-derived DC in type 1 diabetes, whose physiological relevance to endogenous DC is uncertain. Here, we provide the first report characterizing the phenotype and function of endogenous DC subsets in type 1 and type 2 diabetes. We show that DC subsets in each diabetic group exhibit normal properties concerning frequency and activation state, as determined using 4-color flow cytometry of whole blood cells. DC maturation is also intact as confirmed by their efficacious ability to stimulate T cell proliferation in an allogeneic MLR assay. Yet we found that DC are poor producers of IFN-alpha (P < 0.05) in human diabetes. IFN-alpha is a potent antiviral agent and therefore its reduced levels may interfere with T cell-mediated immune responses leading to increased susceptibility and persistence of infections in persons with diabetes.

Myeloid dendritic cells in non-obese diabetic mice have elevated costimulatory and T helper-1-inducing abilities.

Type 1 diabetes (T1D) in the non-obese diabetic (NOD) mouse begins with activation of islet-reactive T helper-1 (Th1) cells by dendritic cells (DCs). Since multiple genetic loci contribute to T1D, we evaluated the hypothesis that NOD DCs possess inherent characteristics that contribute to the autoimmune phenotype. When compared to a representative Th1 (C57BL/6) and Th2 (BALB/C) control strain, in vitro generated NOD myeloid DCs matured normally. Functionally, NOD DCs exhibited higher expression of CD80/86 and IL-12 production during stimulation of nai;ve T cells, even in comparison to C57BL/6 DCs, the prototype strain for vigorous, Th1-biased immunity. These features of NOD DCs translated into aberrantly elevated IFN-gamma synthesis, enhanced T-cell proliferation, and heightened CD69 expression. Further, NOR DCs, from an NOD-related, autoimmune-resistant strain, did not display this hyper-responsiveness, suggesting that these abnormalities are genetic features of NOD DCs that are related to disease pathogenesis. Cumulatively, these results indicate that NOD DCs are inherently biased towards abnormally high costimulation and Th1-induction, two features that would be expected to confer activation and persistence of autoreactive T cells.

Chimerism of murine fetal bone marrow by maternal cells occurs in late gestation and persists into adulthood.

Studies of murine severe combined immune-deficient (scid/scid) fetuses gestating in transgene-tagged immune competent dams have established high frequencies of transplacental trafficking of nucleated maternal cells. Maternal cells first appeared in thymus at gestation day (gd) 12.5 and were present in more than 90% of late gestation fetuses. Morphologically heterogeneous maternal cells were located predominantly in bone marrow and thymus and also occasionally in liver, spleen, and nonlymphoid organs. We have now evaluated maternal cell chimerism in offspring with normal lymphoid development. Genetically normal blastocysts from random-bred CD1 mice were transferred to C57BL/6J- lacZ transgene-tagged ROSA26 females. Serial sectioning of fetuses followed by histochemistry for lacZ-expressing cells was used to comprehensively define organs containing maternal cells. Fetuses, sectioned in their entirety, had no detectable maternal cells before gd 16.5. Morphologically homogenous, nucleated maternal cells were first present in fetal bone marrow cavities at gd 16.5 and were evident in all offspring in later gestation. Postnatally, maternal cells were also present in bone marrow cavities into adulthood, as determined by lacZ histochemistry and PCR amplification of the maternal transgene. The frequency of maternally derived cells in postnatal bone marrow was increased compared with late gestation, and occasionally, maternal cells were detected in postnatal spleen. The normalcy of maternal cell transfer to genetically immune competent progeny and their long-term engraftment is suggestive of a functional role for maternal cells in offspring.