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Mark S. Sands - One of the best experts on this subject based on the ideXlab platform.

  • synergistic effects of treating the spinal cord and brain in cln1 disease
    Proceedings of the National Academy of Sciences of the United States of America, 2017
    Co-Authors: Charles Shyng, Mark S. Sands, Jonathan D. Cooper, Hemanth R Nelvagal, Jaana Tyynela, Joshua T Dearborn, Robert E Schmidt
    Abstract:

    Infantile neuronal ceroid lipofuscinosis (INCL, or CLN1 disease) is an inherited neurodegenerative storage disorder caused by a deficiency of the lysosomal enzyme palmitoyl protein thioesterase 1 (PPT1). It was widely believed that the pathology associated with INCL was limited to the brain, but we have now found unexpectedly profound pathology in the human INCL spinal cord. Similar pathological changes also occur at every level of the spinal cord of PPT1-deficient (PPT1-/- ) mice before the onset of neuropathology in the brain. Various forebrain-directed gene therapy approaches have only had limited success in PPT1-/- mice. Targeting the spinal cord via intrathecal administration of an adeno-associated virus (AAV) gene transfer vector significantly prevented pathology and produced significant improvements in life span and motor function in PPT1-/- mice. Surprisingly, forebrain-directed gene therapy resulted in essentially no PPT1 activity in the spinal cord, and vice versa. This leads to a reciprocal pattern of histological correction in the respective tissues when comparing intracranial with intrathecal injections. However, the characteristic pathological features of INCL were almost completely absent in both the brain and spinal cord when intracranial and intrathecal injections of the same AAV vector were combined. Targeting both the brain and spinal cord also produced dramatic and synergistic improvements in motor function with an unprecedented increase in life span. These data show that spinal cord pathology significantly contributes to the clinical progression of INCL and can be effectively targeted therapeutically. This has important implications for the delivery of therapies in INCL, and potentially in other similar disorders.

  • 27 therapeutic efficacy of intracranial and intrathecal aav2 9 PPT1 in infantile batten disease
    Molecular Therapy, 2016
    Co-Authors: Charles Shyng, Jonathan D. Cooper, Hemanth R Nelvagal, Josh T Dearborn, Mark S. Sands
    Abstract:

    Background The neuronal ceroid lipofuscinoses (NCLs) are a group of the most common pediatric neurodegenerative lysosomal storage disorders. Infantile NCL (INCL), caused by a deficiency in the lysosomal enzyme palmitoyl-protein thioesterase-1 (PPT1), is characterized clinically by progressive cognitive and motor decline, profound neurodegeneration and neuroinflammation, and accumulation of autofluorescent storage material (AFSM). Infantile NCL murine model recapitulates the human disease. AAV2/5-PPT1 intracranial (IC) delivery delayed the onset of INCL histopathological markers in the forebrain and cerebellum and improved preclinical outcome measures. However, overall disease progression was only partially corrected suggesting peripheral nervous system involvement. In collaboration with Dr. Jon Cooper (King's College, London), we discovered substantial progressive pathology in the spinal cord: neuronal loss and axon density, significant microgliosis and astrocytosis, and AFSM (Nelvagal H etal, manuscript in prep). These data suggest that the spinal cord could be an important therapeutic target. We hypothesize that IC and intrathecal (IT) gene therapy in combination will significantly improve the lifespan, preclinical outcome measures, and histopathological markers as compared to either therapy alone. Methods We generated five groups (n=10): PPT1-/-, wild type, and PPT1-/- injected with IC, IT, or the combination IC/IT AAV2/9-PPT1. For IC injections, 3-2µl bilateral intracranial injections were performed. For IT injections, one 15µl bolus injection into the lumbar subarachnoid space was performed. The AAV2/9-PPT1 virus was diluted to 1×1012 viral particles/ml. To date, we have collected 3, 5, and 7-month time points for all groups, and have generated a 9-month time point. Samples will be analyzed for PPT1 enzyme activity, AFSM, neuroinflammation and neurohistopathology, spinal cord pathology, and a histochemical stain for PPT1. Lifespan, behavior, and brain weight (gross measure of atrophy) will be analyzed. Significance was determined using a 2-way ANOVA test. Results PPT1-/- mice have a median lifespan of 35.8 weeks and rapid decline in rotarod performance beginning at 5 months. There is a progressive decline in PPT1-/- brain weight beginning at 3 months. IT AAV2/9-PPT1 mice have a median lifespan of 48.4 weeks and have a steady decline in rotarod performance beginning at 7 months. There is a progressive decline in the IT mice brain weight compared to wild-type, reaching significance at 7 months (p<. 001); however, it had significantly less atrophy than PPT1-/- brains until 7 months (p<0.05). IC AAV2/9-PPT1 mice have a median lifespan of 58.5 weeks and a rapid decline in rotarod performance beginning at 9 months. IC AAV2/9-PPT1 mice brain weight are not significantly different than wild-type. To date, at 66 weeks, all IC/IT AAV2/9-PPT1 mice are alive. There is a significant decline in IC/IT mice rotarod performance at 15 months. The IC/IT mice brain weight is not significantly different than wild-type. Data for the enzyme activity, neuroinflammatory markers, histopathology, and histochemical stain will be complete by April 2016. Conclusions To date, these data confirm that targeting the entire CNS will provide a significant step for INCL therapy. The combination therapy significantly increases the lifespan beyond that of an additive benefit. As expected, modifying the gene therapy vector from IC-AAV2/5 to IC-AAV2/9 significantly improved preclinical outcome measures. Lastly, the IT AAV2/9-PPT1 injections suggest that spinal cord disease plays an important role in INCL pathogenesis. These findings could form the basis for an effective therapeutic strategy that incorporates targeting multiple facets of INCL disease.

  • astrocytosis in infantile neuronal ceroid lipofuscinosis friend or foe
    Biochemical Society Transactions, 2014
    Co-Authors: Charles Shyng, Mark S. Sands
    Abstract:

    Infantile neuronal ceroid lipofuscinosis (INCL; infantile Batten disease) is an inherited paediatric neurodegenerative disease. INCL is caused by a deficiency in the lysosomal enzyme palmitoyl-protein thioesterase-1 (PPT1) and is thus classified as a lysosomal storage disease. Pathological examination of both human and murine INCL brains reveals progressive, widespread neuroinflammation. In fact, astrocyte activation appears to be the first histological sign of disease. However, the role of astrocytosis in INCL was poorly understood. The hallmark of astrocyte activation is the up-regulation of intermediate filaments, such as glial fibrillary acidic protein (GFAP) and vimentin. The role of astrocytosis in INCL was studied in a murine model lacking PPT1 and the intermediate filaments GFAP and vimentin (triple-knockout). This murine model of INCL with attenuated astrocytosis had an exacerbated pathological and clinical phenotype. The triple-knockout mouse had a significantly shortened lifespan, and accelerated cellular and humoural neuroinflammatory response compared with the parental PPT1−/− mouse. The data obtained from the triple-knockout mouse strongly suggest that astrocyte activation plays a beneficial role in early INCL disease progression. A more thorough understanding of the glial responses to lysosomal enzyme deficiencies and the accumulation of undergraded substrates will be crucial to developing effective therapeutics. Abbreviations: CNS, central nervous system; GFAP, glial fibrillary acidic protein; INCL, infantile neuronal ceroid lipofuscinosis; LSD, lysosomal storage disorder; NCL, neuronal ceroid lipofuscinosis; PPT1, palmitoyl-protein thioesterase-1

  • synergistic effects of central nervous system directed gene therapy and bone marrow transplantation in the murine model of infantile neuronal ceroid lipofuscinosis
    Annals of Neurology, 2012
    Co-Authors: Shannon L Macauley, Jonathan D. Cooper, Marie S Roberts, Andrew Wong, Francesca Mcsloy, Adarsh S Reddy, Mark S. Sands
    Abstract:

    Objective: Infantile neuronal ceroid lipofuscinosis (INCL) is an inherited childhood neurodegenerative disorder caused by the loss of palmitoyl protein thioesterase-1 (PPT1) activity. Affected children suffer from blindness, epilepsy, motor dysfunction, cognitive decline, and premature death. The PPT1−/− mouse shares the histological and clinical features of INCL. Previous single-therapy approaches using small molecule drugs, gene therapy, or neuronal stem cells resulted in partial histological correction, with minimal improvements in motor function or lifespan. Here, we combined central nervous system (CNS)-directed adeno-associated virus (AAV)2/5-mediated gene therapy with bone marrow transplantation (BMT) in the INCL mouse. Methods: At birth, PPT1−/− and wild-type mice were given either intracranial injections of AAV2/5-PPT1 or bone marrow transplantation, separately as well as in combination. To assess function, we measured rotorod performance monthly as well as lifespan. At terminal time points, we evaluated the therapeutic effects on several INCL-specific parameters, such as cortical thickness, autofluorescent accumulation, and glial activation. Finally, we determined levels of PPT1 enzyme activity and bone marrow engraftment in treated mice. Results: AAV2/5-mediated gene therapy alone resulted in significant histological correction, improved motor function, and increased lifespan. Interestingly, the addition of BMT further increased the lifespan of treated mice and led to dramatic, sustained improvements in motor function. These data are truly striking, given that BMT alone is ineffective, yet it synergizes with CNS-directed gene therapy to dramatically increase efficacy and lifespan. Interpretation: AAV2/5-mediated gene therapy in combination with BMT provides an unprecedented increase in lifespan as well as dramatic improvement on functional and histological parameters. ANN NEUROL 2012;

  • combination small molecule PPT1 mimetic and cns directed gene therapy as a treatment for infantile neuronal ceroid lipofuscinosis
    Journal of Inherited Metabolic Disease, 2012
    Co-Authors: Marie S Roberts, Shannon L Macauley, Jonathan D. Cooper, Andrew Wong, Denis Yilmas, Sarah A Hohm, Mark S. Sands
    Abstract:

    Infantile neuronal ceroid lipofuscinosis (INCL) is a profoundly neurodegenerative disease of children caused by a deficiency in the lysosomal enzyme palmitoyl protein thioesterase-1 (PPT1). There is currently no effective therapy for this invariably fatal disease. To date, preclinical experiments using single treatments have resulted in incremental clinical improvements. Therefore, we determined the efficacy of CNS-directed AAV2/5-mediated gene therapy alone and in combination with the systemic delivery of the lysosomotropic PPT1 mimetic phosphocysteamine. Since CNS-directed gene therapy provides relatively high levels of PPT1 activity to specific regions of the brain, we hypothesized that phosphocysteamine would complement that activity in regions expressing subtherapeutic levels of the enzyme. Results indicate that CNS-directed gene therapy alone provided the greatest improvements in biochemical and histological measures as well as motor function and life span. Phosphocysteamine alone resulted in only minor improvements in motor function and no increase in lifespan. Interestingly, phosphocysteamine did not increase the biochemical and histological response when combined with AAV2/5-mediated gene therapy, but it did result in an additional improvement in motor function. These data suggest that a CNS-directed gene therapy approach provides significant clinical benefit, and the addition of the small molecule PPT1 mimetic can further increase that response.

Jonathan D. Cooper - One of the best experts on this subject based on the ideXlab platform.

  • synergistic effects of treating the spinal cord and brain in cln1 disease
    Proceedings of the National Academy of Sciences of the United States of America, 2017
    Co-Authors: Charles Shyng, Mark S. Sands, Jonathan D. Cooper, Hemanth R Nelvagal, Jaana Tyynela, Joshua T Dearborn, Robert E Schmidt
    Abstract:

    Infantile neuronal ceroid lipofuscinosis (INCL, or CLN1 disease) is an inherited neurodegenerative storage disorder caused by a deficiency of the lysosomal enzyme palmitoyl protein thioesterase 1 (PPT1). It was widely believed that the pathology associated with INCL was limited to the brain, but we have now found unexpectedly profound pathology in the human INCL spinal cord. Similar pathological changes also occur at every level of the spinal cord of PPT1-deficient (PPT1-/- ) mice before the onset of neuropathology in the brain. Various forebrain-directed gene therapy approaches have only had limited success in PPT1-/- mice. Targeting the spinal cord via intrathecal administration of an adeno-associated virus (AAV) gene transfer vector significantly prevented pathology and produced significant improvements in life span and motor function in PPT1-/- mice. Surprisingly, forebrain-directed gene therapy resulted in essentially no PPT1 activity in the spinal cord, and vice versa. This leads to a reciprocal pattern of histological correction in the respective tissues when comparing intracranial with intrathecal injections. However, the characteristic pathological features of INCL were almost completely absent in both the brain and spinal cord when intracranial and intrathecal injections of the same AAV vector were combined. Targeting both the brain and spinal cord also produced dramatic and synergistic improvements in motor function with an unprecedented increase in life span. These data show that spinal cord pathology significantly contributes to the clinical progression of INCL and can be effectively targeted therapeutically. This has important implications for the delivery of therapies in INCL, and potentially in other similar disorders.

  • 27 therapeutic efficacy of intracranial and intrathecal aav2 9 PPT1 in infantile batten disease
    Molecular Therapy, 2016
    Co-Authors: Charles Shyng, Jonathan D. Cooper, Hemanth R Nelvagal, Josh T Dearborn, Mark S. Sands
    Abstract:

    Background The neuronal ceroid lipofuscinoses (NCLs) are a group of the most common pediatric neurodegenerative lysosomal storage disorders. Infantile NCL (INCL), caused by a deficiency in the lysosomal enzyme palmitoyl-protein thioesterase-1 (PPT1), is characterized clinically by progressive cognitive and motor decline, profound neurodegeneration and neuroinflammation, and accumulation of autofluorescent storage material (AFSM). Infantile NCL murine model recapitulates the human disease. AAV2/5-PPT1 intracranial (IC) delivery delayed the onset of INCL histopathological markers in the forebrain and cerebellum and improved preclinical outcome measures. However, overall disease progression was only partially corrected suggesting peripheral nervous system involvement. In collaboration with Dr. Jon Cooper (King's College, London), we discovered substantial progressive pathology in the spinal cord: neuronal loss and axon density, significant microgliosis and astrocytosis, and AFSM (Nelvagal H etal, manuscript in prep). These data suggest that the spinal cord could be an important therapeutic target. We hypothesize that IC and intrathecal (IT) gene therapy in combination will significantly improve the lifespan, preclinical outcome measures, and histopathological markers as compared to either therapy alone. Methods We generated five groups (n=10): PPT1-/-, wild type, and PPT1-/- injected with IC, IT, or the combination IC/IT AAV2/9-PPT1. For IC injections, 3-2µl bilateral intracranial injections were performed. For IT injections, one 15µl bolus injection into the lumbar subarachnoid space was performed. The AAV2/9-PPT1 virus was diluted to 1×1012 viral particles/ml. To date, we have collected 3, 5, and 7-month time points for all groups, and have generated a 9-month time point. Samples will be analyzed for PPT1 enzyme activity, AFSM, neuroinflammation and neurohistopathology, spinal cord pathology, and a histochemical stain for PPT1. Lifespan, behavior, and brain weight (gross measure of atrophy) will be analyzed. Significance was determined using a 2-way ANOVA test. Results PPT1-/- mice have a median lifespan of 35.8 weeks and rapid decline in rotarod performance beginning at 5 months. There is a progressive decline in PPT1-/- brain weight beginning at 3 months. IT AAV2/9-PPT1 mice have a median lifespan of 48.4 weeks and have a steady decline in rotarod performance beginning at 7 months. There is a progressive decline in the IT mice brain weight compared to wild-type, reaching significance at 7 months (p<. 001); however, it had significantly less atrophy than PPT1-/- brains until 7 months (p<0.05). IC AAV2/9-PPT1 mice have a median lifespan of 58.5 weeks and a rapid decline in rotarod performance beginning at 9 months. IC AAV2/9-PPT1 mice brain weight are not significantly different than wild-type. To date, at 66 weeks, all IC/IT AAV2/9-PPT1 mice are alive. There is a significant decline in IC/IT mice rotarod performance at 15 months. The IC/IT mice brain weight is not significantly different than wild-type. Data for the enzyme activity, neuroinflammatory markers, histopathology, and histochemical stain will be complete by April 2016. Conclusions To date, these data confirm that targeting the entire CNS will provide a significant step for INCL therapy. The combination therapy significantly increases the lifespan beyond that of an additive benefit. As expected, modifying the gene therapy vector from IC-AAV2/5 to IC-AAV2/9 significantly improved preclinical outcome measures. Lastly, the IT AAV2/9-PPT1 injections suggest that spinal cord disease plays an important role in INCL pathogenesis. These findings could form the basis for an effective therapeutic strategy that incorporates targeting multiple facets of INCL disease.

  • synergistic effects of central nervous system directed gene therapy and bone marrow transplantation in the murine model of infantile neuronal ceroid lipofuscinosis
    Annals of Neurology, 2012
    Co-Authors: Shannon L Macauley, Jonathan D. Cooper, Marie S Roberts, Andrew Wong, Francesca Mcsloy, Adarsh S Reddy, Mark S. Sands
    Abstract:

    Objective: Infantile neuronal ceroid lipofuscinosis (INCL) is an inherited childhood neurodegenerative disorder caused by the loss of palmitoyl protein thioesterase-1 (PPT1) activity. Affected children suffer from blindness, epilepsy, motor dysfunction, cognitive decline, and premature death. The PPT1−/− mouse shares the histological and clinical features of INCL. Previous single-therapy approaches using small molecule drugs, gene therapy, or neuronal stem cells resulted in partial histological correction, with minimal improvements in motor function or lifespan. Here, we combined central nervous system (CNS)-directed adeno-associated virus (AAV)2/5-mediated gene therapy with bone marrow transplantation (BMT) in the INCL mouse. Methods: At birth, PPT1−/− and wild-type mice were given either intracranial injections of AAV2/5-PPT1 or bone marrow transplantation, separately as well as in combination. To assess function, we measured rotorod performance monthly as well as lifespan. At terminal time points, we evaluated the therapeutic effects on several INCL-specific parameters, such as cortical thickness, autofluorescent accumulation, and glial activation. Finally, we determined levels of PPT1 enzyme activity and bone marrow engraftment in treated mice. Results: AAV2/5-mediated gene therapy alone resulted in significant histological correction, improved motor function, and increased lifespan. Interestingly, the addition of BMT further increased the lifespan of treated mice and led to dramatic, sustained improvements in motor function. These data are truly striking, given that BMT alone is ineffective, yet it synergizes with CNS-directed gene therapy to dramatically increase efficacy and lifespan. Interpretation: AAV2/5-mediated gene therapy in combination with BMT provides an unprecedented increase in lifespan as well as dramatic improvement on functional and histological parameters. ANN NEUROL 2012;

  • combination small molecule PPT1 mimetic and cns directed gene therapy as a treatment for infantile neuronal ceroid lipofuscinosis
    Journal of Inherited Metabolic Disease, 2012
    Co-Authors: Marie S Roberts, Shannon L Macauley, Jonathan D. Cooper, Andrew Wong, Denis Yilmas, Sarah A Hohm, Mark S. Sands
    Abstract:

    Infantile neuronal ceroid lipofuscinosis (INCL) is a profoundly neurodegenerative disease of children caused by a deficiency in the lysosomal enzyme palmitoyl protein thioesterase-1 (PPT1). There is currently no effective therapy for this invariably fatal disease. To date, preclinical experiments using single treatments have resulted in incremental clinical improvements. Therefore, we determined the efficacy of CNS-directed AAV2/5-mediated gene therapy alone and in combination with the systemic delivery of the lysosomotropic PPT1 mimetic phosphocysteamine. Since CNS-directed gene therapy provides relatively high levels of PPT1 activity to specific regions of the brain, we hypothesized that phosphocysteamine would complement that activity in regions expressing subtherapeutic levels of the enzyme. Results indicate that CNS-directed gene therapy alone provided the greatest improvements in biochemical and histological measures as well as motor function and life span. Phosphocysteamine alone resulted in only minor improvements in motor function and no increase in lifespan. Interestingly, phosphocysteamine did not increase the biochemical and histological response when combined with AAV2/5-mediated gene therapy, but it did result in an additional improvement in motor function. These data suggest that a CNS-directed gene therapy approach provides significant clinical benefit, and the addition of the small molecule PPT1 mimetic can further increase that response.

  • neuroprotection of host cells by human central nervous system stem cells in a mouse model of infantile neuronal ceroid lipofuscinosis
    Cell Stem Cell, 2009
    Co-Authors: Stanley Tamaki, Jonathan D. Cooper, Yakop Jacobs, Monika Dohse, Alexandra Capela, Michael J Reitsma, Dongping He, Robert Tushinski, Pavel V Belichenko, Ahmad Salehi
    Abstract:

    SUMMARY Infantile neuronal ceroid lipofuscinosis (INCL) is a fatal neurodegenerative disease caused by a deficiency in the lysosomal enzyme palmitoyl protein thioesterase-1 (PPT1). PPT1 knockout mice display hallmarks of INCL and mimic the human pathology: accumulation of lipofuscin, degeneration of CNS neurons, and a shortened life span. Purified nongenetically modified human CNS stem cells, grown as neurospheres (hCNS-SCns), were transplanted into the brains of immunodeficient PPT1 / mice where they engrafted robustly, migrated extensively, and produced sufficient levels of PPT1 to alter host neuropathology. Grafted mice displayed reduced autofluorescent lipofuscin, significant neuroprotection of host hippocampal and cortical neurons, and delayed loss of motor coordination. Early intervention with cellular transplants of hCNS-SCns into the brains of INCL patients may supply a continuous and long-lasting source of the missing PPT1 and provide some therapeutic benefit through protection of endogenous neurons. These data provide the experimental basis for human clinical trials with these banked hCNS-SCns.

Anil B Mukherjee - One of the best experts on this subject based on the ideXlab platform.

  • cln3 mutations underlying juvenile neuronal ceroid lipofuscinosis cause significantly reduced levels of palmitoyl protein thioesterases 1 PPT1 protein and PPT1 enzyme activity in the lysosome
    Journal of Inherited Metabolic Disease, 2019
    Co-Authors: Abhilash P Appu, Maria B Bagh, Tamal Sadhukhan, Avisek Mondal, Sydney Casey, Anil B Mukherjee
    Abstract:

    Mutations in at least 13 different genes (called CLNs) underlie various forms of neuronal ceroid lipofuscinoses (NCLs), a group of the most common neurodegenerative lysosomal storage diseases. While inactivating mutations in the CLN1 gene, encoding palmitoyl-protein thioesterases-1 (PPT1), cause infantile NCL (INCL), those in the CLN3 gene, encoding a protein of unknown function, underlie juvenile NCL (JNCL). PPT1 depalmitoylates S-palmitoylated proteins (constituents of ceroid) required for their degradation by lysosomal hydrolases and PPT1-deficiency causes lysosomal accumulation of autofluorescent ceroid leading to INCL. Because intracellular accumulation of ceroid is a characteristic of all NCLs, a common pathogenic link for these diseases has been suggested. It has been reported that CLN3-mutations suppress the exit of cation-independent mannose 6-phosphate receptor (CI-M6PR) from the trans Golgi network (TGN). Because CI-M6PR transports soluble proteins such as PPT1 from the TGN to the lysosome, we hypothesized that CLN3-mutations may cause lysosomal PPT1-insufficiency contributing to JNCL pathogenesis. Here, we report that the lysosomes in Cln3-mutant mice, which mimic JNCL, and those in cultured cells from JNCL patients, contain significantly reduced levels of PPT1-protein and PPT1-enzyme activity and progressively accumulate autofluorescent ceroid. Furthermore, in JNCL fibroblasts the V0a1 subunit of v-ATPase, which regulates lysosomal acidification, is mislocalized to the plasma membrane instead of its normal location on lysosomal membrane. This defect dysregulates lysosomal acidification, as we previously reported in Cln1 -/- mice, which mimic INCL. Our findings uncover a previously unrecognized role of CLN3 in lysosomal homeostasis and suggest that CLN3-mutations causing lysosomal PPT1-insuffiiciency may at least in part contribute to JNCL pathogenesis.

  • neuroprotection and lifespan extension in PPT1 mice by ntbuha therapeutic implications for incl
    Nature Neuroscience, 2013
    Co-Authors: Chinmoy Sarkar, Zhongjian Zhang, Shiyong Peng, Goutam Chandra, Aiyi Liu, Anil B Mukherjee
    Abstract:

    Infantile neuronal ceroid lipofuscinosis (INCL) is a devastating childhood neurodegenerative lysosomal storage disease (LSD) that has no effective treatment. It is caused by inactivating mutations in the palmitoyl-protein thioesterase-1 (PPT1) gene. PPT1 deficiency impairs the cleavage of thioester linkage in palmitoylated proteins (constituents of ceroid), preventing degradation by lysosomal hydrolases. Consequently, accumulation of lysosomal ceroid leads to INCL. Thioester linkage is cleaved by nucleophilic attack. Hydroxylamine, a potent nucleophilic cellular metabolite, may have therapeutic potential for INCL, but its toxicity precludes clinical application. We found that a hydroxylamine derivative, N-(tert-Butyl) hydroxylamine (NtBuHA), was non-toxic, cleaved thioester linkage in palmitoylated proteins and mediated lysosomal ceroid depletion in cultured cells from INCL patients. In PPT1(-/-) mice, which mimic INCL, NtBuHA crossed the blood-brain barrier, depleted lysosomal ceroid, suppressed neuronal apoptosis, slowed neurological deterioration and extended lifespan. Our findings provide a proof of concept that thioesterase-mimetic and antioxidant small molecules such as NtBuHA are potential drug targets for thioesterase deficiency diseases such as INCL.

  • the blood brain barrier is disrupted in a mouse model of infantile neuronal ceroid lipofuscinosis amelioration by resveratrol
    Human Molecular Genetics, 2012
    Co-Authors: Arjun Saha, Chinmoy Sarkar, Jeeva Munasinghe, Zhongjian Zhang, Satya P Singh, Shiyong Peng, Eryan Kong, Goutam Chandra, Anil B Mukherjee
    Abstract:

    Disruption of the blood-brain barrier (BBB) is a serious complication frequently encountered in neurodegenerative disorders. Infantile neuronal ceroid lipofuscinosis (INCL) is a devastating childhood neurodegenerative lysosomal storage disorder caused by palmitoyl-protein thioesterase-1 (PPT1) deficiency. It remains unclear whether BBB is disrupted in INCL and if so, what might be the molecular mechanism(s) of this complication. We previously reported that the PPT1-knockout (PPT1-KO) mice that mimic INCL manifest high levels of oxidative stress and neuroinflammation. Recently, it has been reported that CD4+ T-helper 17 (TH17) lymphocytes may mediate BBB disruption and neuroinflammation, although the precise molecular mechanism(s) remain unclear. We sought to determine: (i) whether the BBB is disrupted in PPT1-KO mice, (ii) if so, do TH17-lymphocytes underlie this complication, and (iii) how might TH17 lymphocytes breach the BBB. Here, we report that the BBB is disrupted in PPT1-KO mice and that TH17 lymphocytes producing IL-17A mediate disruption of the BBB by stimulating production of matrix metalloproteinases (MMPs), which degrade the tight junction proteins essential for maintaining BBB integrity. Importantly, dietary supplementation of resveratrol (RSV), a naturally occurring antioxidant/anti-inflammatory polyphenol, markedly reduced the levels of TH17 cells, IL-17A and MMPs, and elevated the levels of tight junction proteins, which improved the BBB integrity in PPT1-KO mice. Intriguingly, we found that RSV suppressed the differentiation of CD4+ T lymphocytes to IL-17A-positive TH17 cells. Our findings uncover a mechanism by which TH17 lymphocytes mediate BBB disruption and suggest that small molecules such as RSV that suppress TH17 differentiation are therapeutic targets for neurodegenerative disorders such as INCL.

  • Omega-3 and omega-6 fatty acids suppress ER- and oxidative-stress in cultured neurons and neuronal progenitor cells from mice lacking PPT1
    Neuroscience letters, 2010
    Co-Authors: Sungjo Kim, Chinmoy Sarkar, Zhongjian Zhang, Arjun Saha, Zhenwen Zhao, Anil B Mukherjee
    Abstract:

    Reactive oxygen species (ROS) damage brain lipids, carbohydrates, proteins, as well as DNA and may contribute to neurodegeneration. We previously reported that ER- and oxidative stress cause neuronal apoptosis in infantile neuronal ceroid lipofuscinosis (INCL), a lethal neurodegenerative storage disease, caused by palmitoyl-protein thioesterase-1 (PPT1) deficiency. Polyunsaturated fatty acids (PUFA) are essential components of cell membrane phospholipids in the brain and excessive ROS may cause oxidative damage of PUFA leading to neuronal death. Using cultured neurons and neuroprogenitor cells from mice lacking PPT1, which mimic INCL, we demonstrate that PPT1-deficient neurons and neuroprogenitor cells contain high levels of ROS, which may cause peroxidation of PUFA and render them incapable of providing protection against oxidative stress. We tested whether treatment of these cells with omega-3 or omega-6 PUFA protects the neurons and neuroprogenitor cells from oxidative stress and suppress apoptosis. We report here that both omega-3 and omega-6 fatty acids protect the PPT1-deficient cells from ER- as well as oxidative stress and suppress apoptosis. Our results suggest that PUFA supplementation may have neuroprotective effects in INCL.

  • palmitoyl protein thioesterase 1 deficiency impairs synaptic vesicle recycling at nerve terminals contributing to neuropathology in humans and mice
    Journal of Clinical Investigation, 2008
    Co-Authors: Sungjo Kim, Zhongjian Zhang, Chinmoy Sarkar, Peichih Tsai, Yiching Lee, Louis Dye, Anil B Mukherjee
    Abstract:

    Neuronal ceroid lipofuscinoses represent the most common childhood neurodegenerative storage disorders. Infantile neuronal ceroid lipofuscinosis (INCL) is caused by palmitoyl protein thioesterase-1 (PPT1) deficiency. Although INCL patients show signs of abnormal neurotransmission, manifested by myoclonus and seizures, the molecular mechanisms by which PPT1 deficiency causes this abnormality remain obscure. Neurotransmission relies on repeated cycles of exo- and endocytosis of the synaptic vesicles (SVs), in which several palmitoylated proteins play critical roles. These proteins facilitate membrane fusion, which is required for neurotransmitter exocytosis, recycling of the fused SV membrane components, and regeneration of fresh vesicles. However, palmitoylated proteins require depalmitoylation for recycling. Using postmortem brain tissues from an INCL patient and tissue from the PPT1-knockout (PPT1-KO) mice that mimic INCL, we report here that PPT1 deficiency caused persistent membrane anchorage of the palmitoylated SV proteins, which hindered the recycling of the vesicle components that normally fuse with the presynaptic plasma membrane during SV exocytosis. Thus, the regeneration of fresh SVs, essential for maintaining the SV pool size at the synapses, was impaired, leading to a progressive loss of readily releasable SVs and abnormal neurotransmission. This abnormality may contribute to INCL neuropathology.

Sandra L Hofmann - One of the best experts on this subject based on the ideXlab platform.

  • Cross-Linked Enzyme Aggregates as Versatile Tool for Enzyme Delivery: Application to Polymeric Nanoparticles
    2018
    Co-Authors: Marianna Galliani, Sandra L Hofmann, Melissa Santi, Ambra Del Grosso, Antonella Cecchettini, Filippo Maria Santorelli, Lucia Angella, Marco Cecchini, Giovanni Signore
    Abstract:

    Polymeric nanoparticles (NPs) represent one of the most promising tools in nanomedicine and have been extensively studied for the delivery of water-insoluble drugs. However, the efficient loading of therapeutic enzymes and proteins in polymer-based nanostructures remains an open challenge. Here, we report a synthesis method for a new enzyme delivery system based on cross-linked enzyme aggregates (CLEAs) encapsulation into poly­(lactide-co-glycolide) (PLGA) NPs. We tested the encapsulation strategy on four enzymes currently investigated for enzyme replacement therapy: palmitoyl protein thioesterase 1 (PPT1; defective in NCL1 disease), galactosylceramidase (GALC; defective in globoid cell leukodystrophy), alpha glucosidase (aGLU; defective in Pompe disease), and beta glucosidase (bGLU; defective in Gaucher’s disease). We demonstrated that our system allows encapsulation of enzymes with excellent activity retention (usually around 60%), thus leading to functional and targeted nanostructures suitable for enzyme delivery. We then demonstrated that CLEA NPs efficiently deliver PPT1 in cultured cells, with almost complete enzyme release occurring in 48 h. Finally, we demonstrated that enzymatic activity is fully recovered in primary NCL1 fibroblasts upon treatment with PPT1 CLEA NPs

  • a mutation in canine PPT1 causes early onset neuronal ceroid lipofuscinosis in a dachshund
    Molecular Genetics and Metabolism, 2010
    Co-Authors: Douglas N Sanders, Sandra L Hofmann, Fabiana H G Farias, Gary S Johnson, Vivian Chiang, James R Cook, Dennis P Obrien, Jui Yun Lu, Martin L Katz
    Abstract:

    The neuronal ceroid lipofuscinoses (NCLs) are lysosomal storage diseases characterized by progressive neurodegeneration and accumulation of autofluorescent storage granules. A 9-month-old Miniature Dachshund presented with NCL-like signs that included disorientation, ataxia, weakness, visual impairment, and behavioral changes. Neurons throughout the CNS contained autofluorescent lysosomal inclusions with granular osmiophilic deposit (GROD) ultrastructure characteristic of classical infantile NCL (INCL). Human INCL is an autosomal recessive disorder that results from mutations in PPT1, a gene that encodes the enzyme palmitoyl protein thioesterase 1 (PPT1; EC 3.1.22). Resequencing of PPT1 from the affected dog revealed that the dog was homozygous for a single nucleotide insertion in exon 8 (PPT1 c.736_737insC), upstream from the His289 active site. Brain tissue from this dog lacked PPT1 activity. The sire and dam of the propositus were heterozygous for the c.736_737insC mutation; whereas, 127 unrelated Dachshunds were homozygous for the wild-type allele. This is the first reported instance of canine NCL caused by a mutation in PPT1.

  • human recombinant palmitoyl protein thioesterase 1 PPT1 for preclinical evaluation of enzyme replacement therapy for infantile neuronal ceroid lipofuscinosis
    Molecular Genetics and Metabolism, 2010
    Co-Authors: Sandra L Hofmann
    Abstract:

    Infantile neuronal ceroid lipofuscinosis (INCL, also known as Haltia-Santavuori disease) is a lysosomal storage disorder of infants and children characterized by blindness, seizures and a progressive neurodegenerative course. Recent clinical trials have involved neural stem cells and gene therapy directed to the central nervous system; however, enzyme replacement therapy has never been addressed. In the current paper, we describe the production of human recombinant PPT1 (the defective enzyme in INCL) by standard methods in Chinese Hamster Ovary (CHO) cells. The enzyme is largely mannose 6-phosphorylated as assessed by mannose 6-phosphate receptor binding (80% bound) and taken up rapidly by immortalized patient lymphoblasts, where clearance of PPT substrates was demonstrated (EC(50) of 0.25 nM after overnight incubation). When injected intravenously into PPT1-deficient mice, the clearance of recombinant human PPT1 from plasma was rapid, with a half-life of 10 min. Most of the injected dose was distributed to the kidney and liver and potentially corrective levels were also observed in heart, lung and spleen. Brain uptake was minimal, as expected based on experience with other intravenously administered lysosomal enzymes. The enzyme may be useful as an adjunct to central nervous system-directed therapies and could be used as a starting point for modifications designed to improve brain delivery.

  • regional and cellular neuropathology in the palmitoyl protein thioesterase 1 null mutant mouse model of infantile neuronal ceroid lipofuscinosis
    Neurobiology of Disease, 2004
    Co-Authors: Ellen Bible, Praveena Gupta, Sandra L Hofmann, Jonathan D. Cooper
    Abstract:

    Abstract Infantile neuronal ceroid lipofuscinosis (INCL) is one of a group of fatal hereditary lysosomal storage disorders. Palmitoyl protein thioesterase 1 null mutant mice (PPT1−/−) now exist that accurately recapitulate many important disease features. The severely affected PPT1−/− mouse CNS exhibited reduced volume of both cortical and subcortical regions, but with sparing of the cerebellum. Pronounced differences existed in the extent of cortical thinning between different regions, due to lamina-specific effects upon neuronal survival. A dramatic reduction in cortical and hippocampal interneuron number was also evident, with different extents of specific interneuron loss depending upon the region and phenotypic marker. These neuronal changes were accompanied by widespread astrocytosis and localized microglial activation in restricted cortical and subcortical regions. This characterization of PPT1−/− mice not only provides defined pathological landmarks for understanding disease pathogenesis, but also provides an invaluable resource for subsequently judging the efficacy of therapeutic strategies.

  • the crystal structure of palmitoyl protein thioesterase 2 ppt2 reveals the basis for divergent substrate specificities of the two lysosomal thioesterases PPT1 and ppt2
    Journal of Biological Chemistry, 2003
    Co-Authors: Guillermo Calero, Praveena Gupta, Sandra L Hofmann, Jon Clardy, Cristina M Nonato, Sagun Tandel, Edward R Biehl
    Abstract:

    Mutations in palmitoyl protein thioesterase-1 (PPT1) have been found to cause the infantile form of neuronal ceroid lipofuscinosis, which is a lysosomal storage disorder characterized by impaired degradation of fatty acid-modified proteins with accumulation of amorphous granular deposits in cortical neurons, leading to mental retardation and death. Palmitoyl protein thioesterase-2 (PPT2) is a second lysosomal hydrolase that shares a 26% identity with PPT1. A previous study had suggested that palmitoyl-CoA was the preferred substrate of PPT2. Furthermore, PPT2 did not hydrolyze palmitate from the several S-palmitoylated protein substrates. Interestingly, PPT2 deficiency in a recent transgenic mouse model is associated with a form of neuronal ceroid lipofuscinosis, suggesting that PPT1 and -2 perform non-redundant roles in lysosomal thioester catabolism. In the current paper, we present the crystal structure of PPT2 at a resolution of 2.7 A. Comparisons of the structures of PPT1 and -2 show very similar architectural features; however, conformational differences in helix α4 lead to a solvent-exposed lipid-binding groove in PPT1. The limited space between two parallel loops (β3-αA and β8-αF) located immediately above the lipid-binding groove in PPT2 restricts the binding of fatty acids with bulky head groups, and this binding groove is significantly larger in PPT1. This structural difference accounts for the ability of PPT2 to hydrolyze an unbranched structure such as palmitoyl-CoA but not palmitoylcysteine or palmitoylated proteins. Furthermore, differences in fatty acid chain length specificity of PPT1 and -2, also reported here, are explained by the structure and may provide a biochemical basis for their non-redundant roles.

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  • enzymatic activity of palmitoyl protein thioesterase 1 in serum from schizophrenia significantly associates with schizophrenia diagnosis scales
    Journal of Cellular and Molecular Medicine, 2019
    Co-Authors: Qianqian Zhang, Zhongjian Zhang, Jingjing Gao, Guanjie Chen, Xuyi Yue, Shiyong Peng
    Abstract:

    Genome-wide association studies have confirmed that schizophrenia is an inheritable multiple-gene mental disorder. Longitudinal studies about depression, first episode psychosis (FEP) and acute psychotic relapse have mostly searched for brain imaging biomarkers and inflammatory markers from the blood. However, to the best of our knowledge, the association between enzymatic activities with diagnosis or prediction of treatment response in people with schizophrenia has barely been validated. Under the Longitudinal Study of National Mental Health Work Plan (2015-2020), we have studied a subsample of approximately 36 individuals from the cohort with data on palmitoyl-protein thioesterase-1 enzymatic activity from FEP and performed a bivariate correlation analysis with psychiatric assessment scores. After adjusting for sex, age, body mass index (BMI) and total serum protein, our data demonstrated that PPT1 enzymatic activity is significantly associated with schizophrenia and its Positive and Negative Syndrome Scale (PANSS) scores. This longitudinal study compared the PPT1 enzymatic activity in FEP schizophrenia patients and healthy volunteers, and the former exhibited a significant 1.5-fold increase in PPT1 enzymatic levels (1.79 mmol/L/h/mL, and 1.18 mmol/L/h/mL; P < 0.05; 95% CI, 2.3-2.9 and 1.4-1.8). The higher PPT1 enzymatic levels in FEP schizophrenia patients were positively associated with larger PANSS scaling scores (r = 0.32, P = 0.0079 for positive scaling; r = 0.41, P = 0.0006 for negative scaling; r = 0.45, P = 0.0001 for general scaling; and r = 0.34, P = 0.0048 for PNASS-S scaling). Higher enzymatic PPT1 in FEP schizophrenia patients is significantly associated with increased PANSS scaling values, indicating more serious rates of developing psychosis. Enzymatic activity of PPT1 may provide an important new view for schizophrenia disorders.

  • neuroprotection and lifespan extension in PPT1 mice by ntbuha therapeutic implications for incl
    Nature Neuroscience, 2013
    Co-Authors: Chinmoy Sarkar, Zhongjian Zhang, Shiyong Peng, Goutam Chandra, Aiyi Liu, Anil B Mukherjee
    Abstract:

    Infantile neuronal ceroid lipofuscinosis (INCL) is a devastating childhood neurodegenerative lysosomal storage disease (LSD) that has no effective treatment. It is caused by inactivating mutations in the palmitoyl-protein thioesterase-1 (PPT1) gene. PPT1 deficiency impairs the cleavage of thioester linkage in palmitoylated proteins (constituents of ceroid), preventing degradation by lysosomal hydrolases. Consequently, accumulation of lysosomal ceroid leads to INCL. Thioester linkage is cleaved by nucleophilic attack. Hydroxylamine, a potent nucleophilic cellular metabolite, may have therapeutic potential for INCL, but its toxicity precludes clinical application. We found that a hydroxylamine derivative, N-(tert-Butyl) hydroxylamine (NtBuHA), was non-toxic, cleaved thioester linkage in palmitoylated proteins and mediated lysosomal ceroid depletion in cultured cells from INCL patients. In PPT1(-/-) mice, which mimic INCL, NtBuHA crossed the blood-brain barrier, depleted lysosomal ceroid, suppressed neuronal apoptosis, slowed neurological deterioration and extended lifespan. Our findings provide a proof of concept that thioesterase-mimetic and antioxidant small molecules such as NtBuHA are potential drug targets for thioesterase deficiency diseases such as INCL.

  • the blood brain barrier is disrupted in a mouse model of infantile neuronal ceroid lipofuscinosis amelioration by resveratrol
    Human Molecular Genetics, 2012
    Co-Authors: Arjun Saha, Chinmoy Sarkar, Jeeva Munasinghe, Zhongjian Zhang, Satya P Singh, Shiyong Peng, Eryan Kong, Goutam Chandra, Anil B Mukherjee
    Abstract:

    Disruption of the blood-brain barrier (BBB) is a serious complication frequently encountered in neurodegenerative disorders. Infantile neuronal ceroid lipofuscinosis (INCL) is a devastating childhood neurodegenerative lysosomal storage disorder caused by palmitoyl-protein thioesterase-1 (PPT1) deficiency. It remains unclear whether BBB is disrupted in INCL and if so, what might be the molecular mechanism(s) of this complication. We previously reported that the PPT1-knockout (PPT1-KO) mice that mimic INCL manifest high levels of oxidative stress and neuroinflammation. Recently, it has been reported that CD4+ T-helper 17 (TH17) lymphocytes may mediate BBB disruption and neuroinflammation, although the precise molecular mechanism(s) remain unclear. We sought to determine: (i) whether the BBB is disrupted in PPT1-KO mice, (ii) if so, do TH17-lymphocytes underlie this complication, and (iii) how might TH17 lymphocytes breach the BBB. Here, we report that the BBB is disrupted in PPT1-KO mice and that TH17 lymphocytes producing IL-17A mediate disruption of the BBB by stimulating production of matrix metalloproteinases (MMPs), which degrade the tight junction proteins essential for maintaining BBB integrity. Importantly, dietary supplementation of resveratrol (RSV), a naturally occurring antioxidant/anti-inflammatory polyphenol, markedly reduced the levels of TH17 cells, IL-17A and MMPs, and elevated the levels of tight junction proteins, which improved the BBB integrity in PPT1-KO mice. Intriguingly, we found that RSV suppressed the differentiation of CD4+ T lymphocytes to IL-17A-positive TH17 cells. Our findings uncover a mechanism by which TH17 lymphocytes mediate BBB disruption and suggest that small molecules such as RSV that suppress TH17 differentiation are therapeutic targets for neurodegenerative disorders such as INCL.

  • Omega-3 and omega-6 fatty acids suppress ER- and oxidative-stress in cultured neurons and neuronal progenitor cells from mice lacking PPT1
    Neuroscience letters, 2010
    Co-Authors: Sungjo Kim, Chinmoy Sarkar, Zhongjian Zhang, Arjun Saha, Zhenwen Zhao, Anil B Mukherjee
    Abstract:

    Reactive oxygen species (ROS) damage brain lipids, carbohydrates, proteins, as well as DNA and may contribute to neurodegeneration. We previously reported that ER- and oxidative stress cause neuronal apoptosis in infantile neuronal ceroid lipofuscinosis (INCL), a lethal neurodegenerative storage disease, caused by palmitoyl-protein thioesterase-1 (PPT1) deficiency. Polyunsaturated fatty acids (PUFA) are essential components of cell membrane phospholipids in the brain and excessive ROS may cause oxidative damage of PUFA leading to neuronal death. Using cultured neurons and neuroprogenitor cells from mice lacking PPT1, which mimic INCL, we demonstrate that PPT1-deficient neurons and neuroprogenitor cells contain high levels of ROS, which may cause peroxidation of PUFA and render them incapable of providing protection against oxidative stress. We tested whether treatment of these cells with omega-3 or omega-6 PUFA protects the neurons and neuroprogenitor cells from oxidative stress and suppress apoptosis. We report here that both omega-3 and omega-6 fatty acids protect the PPT1-deficient cells from ER- as well as oxidative stress and suppress apoptosis. Our results suggest that PUFA supplementation may have neuroprotective effects in INCL.

  • palmitoyl protein thioesterase 1 deficiency impairs synaptic vesicle recycling at nerve terminals contributing to neuropathology in humans and mice
    Journal of Clinical Investigation, 2008
    Co-Authors: Sungjo Kim, Zhongjian Zhang, Chinmoy Sarkar, Peichih Tsai, Yiching Lee, Louis Dye, Anil B Mukherjee
    Abstract:

    Neuronal ceroid lipofuscinoses represent the most common childhood neurodegenerative storage disorders. Infantile neuronal ceroid lipofuscinosis (INCL) is caused by palmitoyl protein thioesterase-1 (PPT1) deficiency. Although INCL patients show signs of abnormal neurotransmission, manifested by myoclonus and seizures, the molecular mechanisms by which PPT1 deficiency causes this abnormality remain obscure. Neurotransmission relies on repeated cycles of exo- and endocytosis of the synaptic vesicles (SVs), in which several palmitoylated proteins play critical roles. These proteins facilitate membrane fusion, which is required for neurotransmitter exocytosis, recycling of the fused SV membrane components, and regeneration of fresh vesicles. However, palmitoylated proteins require depalmitoylation for recycling. Using postmortem brain tissues from an INCL patient and tissue from the PPT1-knockout (PPT1-KO) mice that mimic INCL, we report here that PPT1 deficiency caused persistent membrane anchorage of the palmitoylated SV proteins, which hindered the recycling of the vesicle components that normally fuse with the presynaptic plasma membrane during SV exocytosis. Thus, the regeneration of fresh SVs, essential for maintaining the SV pool size at the synapses, was impaired, leading to a progressive loss of readily releasable SVs and abnormal neurotransmission. This abnormality may contribute to INCL neuropathology.