Daniel Larocque

Toll-like receptor 4 stimulation with the detoxified ligand monophosphoryl lipid A improves Alzheimer’s disease-related pathology

Alzheimer’s disease (AD) is the most common cause of dementia worldwide. The pathogenesis of this neurodegenerative disease, currently without curative treatment, is associated with the accumulation of amyloid β (Aβ) in brain parenchyma and cerebral vasculature. AD patients are unable to clear this toxic peptide, leading to Aβ accumulation in their brains and, presumably, the pathology associated with this devastating disease. Compounds that stimulate the immune system to clear Aβ may therefore have great therapeutic potential in AD patients. Monophosphoryl lipid A (MPL) is an LPS-derived Toll-like receptor 4 agonist that exhibits unique immunomodulatory properties at doses that are nonpyrogenic. We show here that repeated systemic injections of MPL, but not LPS, significantly improved AD-related pathology in APPswe/PS1 mice. MPL treatment led to a significant reduction in Aβ load in the brain of these mice, as well as enhanced cognitive function. MPL induced a potent phagocytic response by microglia while triggering a moderate inflammatory reaction. Our data suggest that the Toll-like receptor 4 agonist MPL may be a treatment for AD.

Combined Flt3L/TK Gene Therapy Induces Immunological Surveillance Which Mediates an Immune Response Against a Surrogate Brain Tumor Neoantigen

Glioblastoma multiforme (GBM) is a primary brain tumor with a median survival of 14.6 months postdiagnosis. The infiltrative nature of GBM prevents complete resection and residual brain tumor cells give rise to recurrent GBM, a hallmark of this disease. Recurrent GBMs are known to harbor numerous mutations/gene rearrangements when compared to the primary tumor, which leads to the potential expression of novel proteins that could serve as tumor neoantigens. We have developed a combined immune-based gene therapeutic approach for GBM using adenoviral (Ads) mediated gene delivery of Herpes Simplex Virus Type 1-thymidine kinase (TK) into the tumor mass to induce tumor cells death combined with an adenovirus expressing fms-like tyrosine kinase 3 ligand (Flt3L) to recruit dendritic cells (DCs) into the tumor microenvironment. This leads to the induction of specific anti-brain tumor immunity and immunological memory. In a model of GBM recurrence, we demonstrate that Flt3L/TK mediated immunological memory is capable of recognizing brain tumor neoantigens absent from the original treated tumor. These data demonstrate that the Flt3L/TK gene therapeutic approach can induce systemic immunological memory capable of recognizing a brain tumor neoantigen in a model of recurrent GBM.Glioblastoma multiforme (GBM) is a primary brain tumor with a median survival of 14.6 months postdiagnosis. The infiltrative nature of GBM prevents complete resection and residual brain tumor cells give rise to recurrent GBM, a hallmark of this disease. Recurrent GBMs are known to harbor numerous mutations/gene rearrangements when compared to the primary tumor, which leads to the potential expression of novel proteins that could serve as tumor neoantigens. We have developed a combined immune-based gene therapeutic approach for GBM using adenoviral (Ads) mediated gene delivery of Herpes Simplex Virus Type 1-thymidine kinase (TK) into the tumor mass to induce tumor cells death combined with an adenovirus expressing fms-like tyrosine kinase 3 ligand (Flt3L) to recruit dendritic cells (DCs) into the tumor microenvironment. This leads to the induction of specific anti-brain tumor immunity and immunological memory. In a model of GBM recurrence, we demonstrate that Flt3L/TK mediated immunological memory is capable of recognizing brain tumor neoantigens absent from the original treated tumor. These data demonstrate that the Flt3L/TK gene therapeutic approach can induce systemic immunological memory capable of recognizing a brain tumor neoantigen in a model of recurrent GBM.

Immunization against the transgene but not the TetON switch reduces expression from gutless adenoviral vectors in the brain.

Immune responses against vectors or encoded transgenes can impose limitations on gene therapy. We demonstrated that tetracycline-regulated high-capacity adenoviral vectors (HC-Ads) sustain regulated transgene expression in the brain even in the presence of systemic pre-existing immune responses against adenoviruses. In this study we assessed whether systemic pre-existing immune responses against the transgene products, i.e., beta-Gal or the tetracycline-dependent (TetON) regulatory transcription factors (rtTA2(S)M2 and the tTS(Kid)), affect transgene expression levels and the safety profile of HC-Ads in the brain. We pre-immunized mice with plasmids encoding the TetON switch expressing rtTA2(S)M2 and the tTS(Kid) or beta-Gal. HC-Ads expressing beta-Gal under the control of the TetON switch were then injected into the striatum. We assessed levels and distribution of beta-Gal expression, and evaluated local inflammation and neuropathological changes. We found that systemic immunity against beta-Gal, but not against the TetON switch, led to inflammation and reduction of transgene expression in the striatum. Therefore, the regulatory TetON switch appears to be safe to use, and capable of sustaining transgene expression in the brain even in the presence of an immune response against its components. Systemic immunity against the transgene had the effect of curtailing its expression, thereby affecting the efficacy and safety of gene delivery to the brain. This factor should be considered when developing gene therapies for neurological use.

High-Capacity Adenovirus Vector-Mediated Anti-Glioma Gene Therapy in the Presence of Systemic Antiadenovirus Immunity

ABSTRACT Gene therapy is proposed as a novel therapeutic strategy for treating glioblastoma multiforme (GBM), a devastating brain cancer. In the clinic, antivector immune responses pose formidable challenges. Herein we demonstrate that high-capacity adenovirus vectors (HC-Ads) carrying the conditional cytotoxic gene herpes simplex virus type 1-thymidine kinase (TK) induce tumor regression and long-term survival in an intracranial glioma model, even in the presence of systemic antiadenovirus immunity, as could be encountered in patients. First-generation Ad-TK failed to elicit tumor regression in this model. These results pave the way for implementing HC-Ad-TK-mediated gene therapy as a powerful adjuvant for treating GBM.

Exogenous fms-like tyrosine kinase 3 ligand overrides brain immune privilege and facilitates recognition of a neo-antigen without causing autoimmune neuropathology

Soluble antigens diffuse out of the brain and can thus stimulate a systemic immune response, whereas particulate antigens (from infectious agents or tumor cells) remain within brain tissue, thus failing to stimulate a systemic immune response. Immune privilege describes how the immune system responds to particulate antigens localized selectively within the brain parenchyma. We believe this immune privilege is caused by the absence of antigen presenting dendritic cells from the brain. We tested the prediction that expression of fms-like tyrosine kinase ligand 3 (Flt3L) in the brain will recruit dendritic cells and induce a systemic immune response against exogenous influenza hemagglutinin in BALB/c mice. Coexpression of Flt3L with HA in the brain parenchyma induced a robust systemic anti-HA immune response, and a small response against myelin basic protein and proteolipid protein epitopes. Depletion of CD4+CD25+ regulatory T cells (Tregs) enhanced both responses. To investigate the autoimmune impact of these immune responses, we characterized the neuropathological and behavioral consequences of intraparenchymal injections of Flt3L and HA in BALB/c and C57BL/6 mice. T cell infiltration in the forebrain was time and strain dependent, and increased in animals treated with Flt3L and depleted of Tregs; however, we failed to detect widespread defects in myelination throughout the forebrain or spinal cord. Results of behavioral tests were all normal. These results demonstrate that Flt3L overcomes the brains immune privilege, and supports the clinical development of Flt3L as an adjuvant to stimulate clinically effective immune responses against brain neo-antigens, for example, those associated with brain tumors.

Immunology and the Central Nervous System

We are pleased to announce the publication of this special issue in the journal “Clinical and Developmental Immunology.” We are happy to perceive the growing interest, from a wide range of scientists, regarding the peculiar intercommunication between the immune system and the central nervous system (CNS). We have finally reached a balanced compilation of papers that we collect in this special issue highlighting recent fundamental advances in our understanding of brain immunology with an emphasis on new therapeutic targets covering such emerging topics as chemical suppression of glial activation, inflammation following acute demylination, Notch signalling as a potential therapeutic target in EAE and a link between neuroinflammatory signaling and reproduction. n nThe idea that the CNS is an immune-privileged site is gradually vanishing. However, increasing evidence shows that the relation between the CNS and the immune system is special and in many aspects, different from the rest of the organs and tissues [1]. n nOne of the peculiarities of the CNS is the presence of glial cells, which are the initial responders to brain injuries and degenerative processes [2]. Glial cells, especially microglia, get locally activated in the damaged brain areas and are able to induce the recruitment of scavenger blood cells, such as monocytes and lymphocytes to injured sites, contributing to the inflammatory reaction [3, 4]. In our special issue, R. A. Taylor and L. H. Sansing, describe, in a comprehensive review, the role of microglial cells in ischemic stroke and intracerebral hemorrhage. Importantly, authors highlight the distinct phenotypes of microglia, M1-inflammatory, and M2-repairing microglia, which express different surface molecules and releasing factors. They propose that understanding the mechanism of this switching phenotype will be crucial for future therapeutic purposes. In line with this, E. Assi et al., review the recent literature about the role of microglia in inflammatory signaling cascades in brain pathology, but focusing their paper on the role of sphingolipids in the inflammatory reaction, which is proposed as a potential target to control glial-mediated neuroinflammation. n nIn this context, it is becoming more evident that glial activation is a critical event that should be targeted to avert inflammation in the CNS. B. Rocamonde et al. describe, in an original study, using a rat model of brain injury, that the reduction of glial activation by lipoic acid underlies the restorative effects in the brain. F. Cloutier et al. suggest in an interesting paper that the role of microglia and astrocytes during spinal cord injury and repair may be different depending on the scenario of CNS damage. Authors report, using a rat model of acute demyelination in dorsal funiculus, how immunological using anti GalC demyelination triggers macrophage/microglial cells activation in comparison of a stab injury. Interestingly, in their model of axon regeneration, the participation of astrocytes is limited, whereas microglia and infiltrated macrophages have a prominent role, which emphasize the potential of targeting microglial cells for therapeutic purposes. n nFrom this perspective, papers on CNS autoimmunity, MS and MS experimental models have largely contributed to our special issue. We have compiled extensive reviews and original papers about the initial immune-pathogenesis of MS as well as the immunology and oxidative stress underlying the disease. R. Bassil et al. provide a comprehensive reassessment on the important function of Notch signaling in the activation of T helper cells in experimental autoimmune encephalitis (EAE). They propose targeting Notch signaling as a potential strategy for MS therapy, but they also report the limitations of this approach due to the wide range of functions of Notch signaling, suggesting interesting new research avenues. Hernandez-N. Y. Pedro et al. discuss the innate immune responses occurring during the immune-pathogenesis of MS, which are hypothesized as a critical trigger of the chronic inflammatory response. In line with this, G. G. Ortiz et al. describe that oxidative stress and inflammation are important elements in the self-perpetuation inflammatory/immune cycle of MS. These pathways may contain targets that could be promising to avert the inflammatory responses in MS. Concluding this MS section from a practical point of view, S. F. Goncalves Zorzella-Pezavento et al contribute to settle down the potential controversy regarding the tuberculosis (TB) vaccination protocols, suggesting that TB vaccination does not trigger or worsen EAE pathology. n nImmunotherapy is also an important topic covered by this special issue. In the CNS, T-lymphocytes infiltrate into the inflamed brain parenchyma and the manipulation of different T-cell subpopulations may have beneficial effect on neurodegenerative disorders and diverse brain injuries. T cell infiltration is observed in many neurodegenerative diseases including Parkinsons and Alzheimers disease and we are getting to know more aspects on how T-cells may affect neurodegeneration [5, 6]. Importantly, immunotherapy based on targeting T cells (Tregs and Th17 cells) for autoimmunity disorders and cancer is becoming a rising field of investigation [7]. However, the precise lymphocyte function in the damaged brain parenchyma is still poorly understood. S. Chen et al. propose, in a completed review, that the modulation of Tregs may have a favorable outcome for stroke patients. This is based on the fact that Tregs downregulates the excessive brain immune-reaction, which could be favorable for tissue restoration. In line with this, M. S. Zaborowski and Michalak describe how the appearance of paraneoplastic neurological syndromes may be linked with the certain antitumor responses triggered by T cells. Thus, authors suggest that the manipulation of specific subpopulation of T cells, pondering the balance between Tregs and CTLs through immunotherapy, could be beneficial for these neurological disorders. n nIn this context of immunotherapy, the search for antigens that may trigger autoimmune responses in the brain is a crucial field of research. The review by A. Seppanen proposes collagen XVII as a candidate antigen. Autoantibodies against different domains of collagen XVII are found elevated in serum of patients within a wide range of neurological disorders as well as in skin autoimmune disorders, suggesting that an autoimmune response may be the common trigger of those disorders. On the other hand, increasing evidence indicates that immune system dysfunction may also be underlying a number of psychiatric disorders. K. Pathmanandavel et al. hypothesize in their thought-provoking review that some neuronal autoantibodies may be linked with psychiatric symptoms and could open new immunomodulatory approaches for neuropsychiatric disorders in the near future. n nWe scarcely know the factors that trigger CNS diseases. It is thought that genetic and environmental factors, which include the exposure to chemical compounds or particular pathogens, may contribute to the appearance of these disorders. T. T. Win-Shwe et al. discuss an interesting point regarding the immune-related inflammation that environmental volatile compounds may induce. The review hypothesizes how environmental volatile elements may affect respiratory and immune system, having neurological consequences for the population. On the other hand, T. Hautala et al. report in our special issue how viruses, in this case Puumala virus, a northern European type of Hantavirus, are important entities that may seriously affect the CNS. Other pathogen infections, such as Trypanosoma brucei, which causes sleeping sickness, may also cause severe brain damage. Interestingly, D. N. Maranga et al., members of a research group based in Kenya, present in our special issue a research study performed in few monkeys showing that the increase of the proinflammatory cytokine IL-6 in spinal fluid may be a reliable marker to manage human African trypanosomiasis in areas where the diagnostic tools are very limited and derived neurological consequences are frequent. Lastly, A. P. Herman et al. describe how the inflammatory-mediated release of cytokines, such as IL-1β, may have consequences in the hypothalamic neurons affecting the reproductive system. This highlights that it is also important to consider the effect that neuroimmune interactions and neuroinflammation may have in many other systems. n nWe hope this collection of papers is helpful for readers to understand better the peculiar relationship between the immune system and CNS and may be a stimulus to continue the research on this complex field. n n nCarlos Barcia n nJames Curtin n nJeffrey Zirger n nDaniel Larocque