Johns Hopkins Drug Discovery - Project - Neutral Sphingomyelinase 2
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Neutral Sphingomyelinase 2

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The objective of Johns Hopkins Drug Discovery’s neutral sphinghomyelinase 2 (nSmase2) project is to optimize a small molecule inhibitor for the treatment of Alzheimer’s disease (AD). NSMase2 catalyzes the hydrolysis of sphinghomyelin (SM) to phosphorylcholine and ceramide. Production of ceramide through nSMase2 upregulation has been associated with diverse cellular processes ranging from apoptosis (1)  to modulation of synaptic plasticity (2) to manufacturing of ceramide-rich exosomes (3). While transient nSMase2 upregulation is part of normal brain functioning, chronic upregulation of the enzyme has been implicated in the pathogenesis of various neurodegenerative diseases associated with excess ceramide including Alzheimer’s disease (AD) (4-9), HIV-associated neurocognitive disorders (HAND) (1, 10), multiple sclerosis (MS) (11) and amyotrophic lateral sclerosis (ALS) (12). NSMase2 is predominantly expressed in the brain and it is this isoform, rather than nSMase 1 or 3, that has been implicated in neurodegenerative disease (13, 14). Post mortem AD brains exhibit abnormal increases in endogenous ceramide (5, 8, 15) and ceramide has been shown to be an integral component of exosomes (3). Recent experimental results suggest that elevation of ceramide-rich exosomes in the brain is involved in AD progression (16, 17),  that these exosomes are involved in tau propagation upon secretion from activated microglia (16) and that inhibition of exosome manufacturing through pharmacological inhibition of nSMase2 prevents tau propagation (16). In another recent, independent study using an AD mouse model, ceramide-enriched exosomes exacerbated AD pathology and cognitive deficits while nSMase2 deficiency ameliorated AD pathology and improved cognition (18). The results provide a compelling rationale for the potential use of nSMase2 inhibitors for the treatment of AD.

 

While nSmase2 has emerged as an important player in AD etiology, the current armamentarium of nSMase2 inhibitors is inadequate to develop potential treatments. Currently available inhibitors have many limitations including low potency (IC50’s in µM level), poor solubility, no brain penetration and their chemical structures have not been amenable to optimization into drug-like molecules (19, 20). In order to address these limitations, we screened several chemical libraries (~ 500,000 compounds) which led to the identification of several hits belonging to different chemical series. One of these hits was significantly improved for potency and selectivity and we now have systematically synthesized several analogs with IC50 < 100 nM that are being further optimized for microsomal stability and brain penetration. Selected optimized compounds will be evaluated in the recently published AD model (16), as well as characterized in models of HAND (2, 10) and multiple sclerosis (21).

 

References

 

  1. Jana, A., and Pahan, K. (2004) Human immunodeficiency virus type 1 gp120 induces apoptosis in human primary neurons through redox-regulated activation of neutral sphingomyelinase, J Neurosci 24, 9531-9540.
  2. Wheeler, D., Knapp, E., Bandaru, V. V., Wang, Y., Knorr, D., Poirier, C., Mattson, M. P., Geiger, J. D., and Haughey, N. J. (2009) Tumor necrosis factor-alpha-induced neutral sphingomyelinase-2 modulates synaptic plasticity by controlling the membrane insertion of NMDA receptors, J Neurochem 109, 1237-1249.
  3. Trajkovic, K., Hsu, C., Chiantia, S., Rajendran, L., Wenzel, D., Wieland, F., Schwille, P., Brugger, B., and Simons, M. (2008) Ceramide triggers budding of exosome vesicles into multivesicular endosomes, Science 319, 1244-1247.
  4. Alessenko, A. V., Bugrova, A. E., and Dudnik, L. B. (2004) Connection of lipid peroxide oxidation with the sphingomyelin pathway in the development of Alzheimer’s disease, Biochem Soc Trans 32, 144-146.
  5. Filippov, V., Song, M. A., Zhang, K., Vinters, H. V., Tung, S., Kirsch, W. M., Yang, J., and Duerksen-Hughes, P. J. (2012) Increased ceramide in brains with Alzheimer’s and other neurodegenerative diseases, J Alzheimers Dis 29, 537-547.
  6. Marks, N., Berg, M. J., and Saito, M. (2008) Glucosylceramide synthase decrease in frontal cortex of Alzheimer brain correlates with abnormal increase in endogenous ceramides: consequences to morphology and viability on enzyme suppression in cultured primary neurons, Brain Res 1191, 136-147.
  7. Mielke, M. M., Haughey, N. J., Bandaru, V. V., Schech, S., Carrick, R., Carlson, M. C., Mori, S., Miller, M. I., Ceritoglu, C., Brown, T., Albert, M., and Lyketsos, C. G. (2010) Plasma ceramides are altered in mild cognitive impairment and predict cognitive decline and hippocampal volume loss, Alzheimers Dement 6, 378-385.
  8. Satoi, H., Tomimoto, H., Ohtani, R., Kitano, T., Kondo, T., Watanabe, M., Oka, N., Akiguchi, I., Furuya, S., Hirabayashi, Y., and Okazaki, T. (2005) Astroglial expression of ceramide in Alzheimer’s disease brains: a role during neuronal apoptosis, Neuroscience 130, 657-666.
  9. Wang, G., Silva, J., Dasgupta, S., and Bieberich, E. (2008) Long-chain ceramide is elevated in presenilin 1 (PS1M146V) mouse brain and induces apoptosis in PS1 astrocytes, Glia 56, 449-456.
  10. Haughey, N. J., Cutler, R. G., Tamara, A., McArthur, J. C., Vargas, D. L., Pardo, C. A., Turchan, J., Nath, A., and Mattson, M. P. (2004) Perturbation of sphingolipid metabolism and ceramide production in HIV-dementia, Ann Neurol 55, 257-267.
  11. Jana, A., and Pahan, K. (2010) Sphingolipids in multiple sclerosis, Neuromolecular Med 12, 351-361.
  12. Cutler, R. G., Pedersen, W. A., Camandola, S., Rothstein, J. D., and Mattson, M. P. (2002) Evidence that accumulation of ceramides and cholesterol esters mediates oxidative stress-induced death of motor neurons in amyotrophic lateral sclerosis, Ann Neurol 52, 448-457.
  13. Wu, B. X., Clarke, C. J., and Hannun, Y. A. (2010) Mammalian neutral sphingomyelinases: regulation and roles in cell signaling responses, Neuromolecular Med 12, 320-330.
  14. Horres, C. R., and Hannun, Y. A. (2012) The roles of neutral sphingomyelinases in neurological pathologies, Neurochem Res 37, 1137-1149.
  15. Mielke, M. M., Bandaru, V. V., McArthur, J. C., Chu, M., and Haughey, N. J. (2010) Disturbance in cerebral spinal fluid sphingolipid content is associated with memory impairment in subjects infected with the human immunodeficiency virus, J Neurovirol 16, 445-456.
  16. Asai, H., Ikezu, S., Tsunoda, S., Medalla, M., Luebke, J., Haydar, T., Wolozin, B., Butovsky, O., Kugler, S., and Ikezu, T. (2015) Depletion of microglia and inhibition of exosome synthesis halt tau propagation, Nat Neurosci 18, 1584-1593.
  17. Joshi, P., Benussi, L., Furlan, R., Ghidoni, R., and Verderio, C. (2015) Extracellular vesicles in Alzheimer’s disease: friends or foes? Focus on abeta-vesicle interaction, Int J Mol Sci 16, 4800-4813.
  18. Dinkins, M. B., Enasko, J., Hernandez, C., Wang, G., Kong, J., Helwa, I., Liu, Y., Terry, A. V., Jr., and Bieberich, E. (2016) Neutral Sphingomyelinase-2 Deficiency Ameliorates Alzheimer’s Disease Pathology and Improves Cognition in the 5XFAD Mouse, J Neurosci 36, 8653-8667.
  19. Luberto, C., Hassler, D. F., Signorelli, P., Okamoto, Y., Sawai, H., Boros, E., Hazen-Martin, D. J., Obeid, L. M., Hannun, Y. A., and Smith, G. K. (2002) Inhibition of tumor necrosis factor-induced cell death in MCF7 by a novel inhibitor of neutral sphingomyelinase, J Biol Chem 277, 41128-41139.
  20. Rojas, C., Frazier, S. T., Flanary, J., and Slusher, B. S. (2002) Kinetics and inhibition of glutamate carboxypeptidase II using a microplate assay, Anal Biochem 310, 50-54.
  21. Matsushima, G. K., and Morell, P. (2001) The neurotoxicant, cuprizone, as a model to study demyelination and remyelination in the central nervous system, Brain Pathol 11, 107-116.
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