In a recent study we found that cerebrospinal fluids (CSFs) from amyotrophic lateral sclerosis (ALS) patients caused 20-30% loss of cell viability in primary cultures of rat embryo motor cortex neurons. We also found that the antioxidant resveratrol protected against such damaging effects and that, surprisingly, riluzole antagonized its protecting effects. Here we have extended this study to the interactions of riluzole with 3 other recognized neuroprotective agents, namely memantine, minocycline and lithium. We found: (1) by itself riluzole exerted neurotoxic effects at concentrations of 3-30 µM; this cell damage was similar to that elicited by 30 µM glutamate and a 10% dilution of ALS/CSF; (2) memantine (0.1-30 µM), minocycline (0.03-1 µM) and lithium (1-80 µg/ml) afforded 10-30% protection against ALS/CSF-elicited neurotoxicity, and (3) at 1-10 µM, riluzole antagonized the protection afforded by the 3 agents. These results strongly support the view that at the riluzole concentrations reached in the brain of patients, the neurotoxic effects of this drug could be masking the potential neuroprotective actions of new compounds being tested in clinical trials. Therefore, in the light of the present results, the inclusion of a group of patients free of riluzole treatment may be mandatory in future clinical trials performed in ALS patients with novel neuroprotective compounds.

Keywords:
Memantine, Minocycline, Lithium, Amyotrophic lateral sclerosis, Neurotoxicity, Riluzole
1.
Bensimon G, Lacomblez L, Meininger V: A controlled trial of riluzole in amyotrophic lateral sclerosis. ALS/Riluzole Study Group. N Engl J Med 1994;330:585-591.
2.
Lacomblez L, Bensimon G, Leigh PN, Guillet P, Meininger V: Dose-ranging study of riluzole in amyotrophic lateral sclerosis. Amyotrophic Lateral Sclerosis/Riluzole Study Group II. Lancet 1996;347:1425-1431.
3.
Wang R, Zhang D: Memantine prolongs survival in an amyotrophic lateral sclerosis mouse model. Eur J Neurosci 2005;22:2376-2380.
4.
Joo IS, Hwang DH, Seok JI, Shin SK, Kim SU: Oral administration of memantine prolongs survival in a transgenic mouse model of amyotrophic lateral sclerosis. J Clin Neurol 2007;3:181-186.
5.
De Carvalho M, Pinto S, Costa J, Evangelista T, Ohana B, Pinto A: A randomized, placebo-controlled trial of memantine for functional disability in amyotrophic lateral sclerosis. Amyotroph Lateral Scler 2010;11:456-460.
6.
Kriz J, Nguyen MD, Julien JP: Minocycline slows disease progression in a mouse model of amyotrophic lateral sclerosis. Neurobiol Dis 2002;10:268-278.
7.
Van Den Bosch L, Tilkin P, Lemmens G, Robberecht W: Minocycline delays disease onset and mortality in a transgenic model of ALS. Neuroreport 2002;13:1067-1070.
8.
Zhu S, Stavrovskaya IG, Drozda M, et al: Minocycline inhibits cytochrome c release and delays progression of amyotrophic lateral sclerosis in mice. Nature 2002;417:74-78.
9.
Zhang W, Narayanan M, Friedlander RM: Additive neuroprotective effects of minocycline with creatine in a mouse model of ALS. Ann Neurol 2003;53:267-270.
10.
Kriz J, Gowing G, Julien JP: Efficient three-drug cocktail for disease induced by mutant superoxide dismutase. Ann Neurol 2003;53:429-436.
11.
Gordon PH, Moore DH, Gelinas DF, et al: Placebo-controlled phase I/II studies of minocycline in amyotrophic lateral sclerosis. Neurology 2004;62:1845-1847.
12.
Pontieri FE, Ricci A, Pellicano C, Benincasa D, Buttarelli FR: Minocycline in amyotrophic lateral sclerosis: a pilot study. Neurol Sci 2005;26:285-287.
13.
Gordon PH, Moore DH, Miller RG, et al: Efficacy of minocycline in patients with amyotrophic lateral sclerosis: a phase III randomised trial. Lancet Neurol 2007;6:1045-1053.
14.
Shin JH, Cho SI, Lim HR, et al: Concurrent administration of Neu2000 and lithium produces marked improvement of motor neuron survival, motor function, and mortality in a mouse model of amyotrophic lateral sclerosis. Mol Pharmacol 2007;71:965-975.
15.
Feng HL, Leng Y, Ma CH, Zhang J, Ren M, Chuang DM: Combined lithium and valproate treatment delays disease onset, reduces neurological deficits and prolongs survival in an amyotrophic lateral sclerosis mouse model. Neuroscience 2008;155:567-572.
16.
Fornai F, Longone P, Cafaro L, et al: Lithium delays progression of amyotrophic lateral sclerosis. Proc Natl Acad Sci USA 2008;105:2052-2057.
17.
Pizzasegola C, Caron I, Daleno C, et al: Treatment with lithium carbonate does not improve disease progression in two different strains of SOD1 mutant mice. Amyotroph Lateral Scler 2009;10:221-228.
18.
Gill A, Kidd J, Vieira F, Thompson K, Perrin S: No benefit from chronic lithium dosing in a sibling-matched, gender balanced, investigator-blinded trial using a standard mouse model of familial ALS. PLoS One 2009;4:e6489.
19.
Chio A, Borghero G, Calvo A, et al: Lithium carbonate in amyotrophic lateral sclerosis: lack of efficacy in a dose-finding trial. Neurology 2010;75:619-625.
20.
Verstraete E, Veldink JH, Huisman MH, et al: Lithium lacks effect on survival in amyotrophic lateral sclerosis: a phase IIb randomised sequential trial. J Neurol Neurosurg Psychiatry 2012;83:557-564.
21.
Terro F, Lesort M, Dussartre C, Barthe D, Hugon J: Phosphorylated neurofilament expression and resistance to kainate toxicity. Brain Res Bull 1996;41:231-235.
22.
Shaw PJ, Eggett CJ: Molecular factors underlying selective vulnerability of motor neurons to neurodegeneration in amyotrophic lateral sclerosis. J Neurol 2000;247(suppl 1):I17-I27.
23.
Shahani N, Gourie-Devi M, Nalini A, Raju TR: Cyclophosphamide attenuates the degenerative changes induced by CSF from patients with amyotrophic lateral sclerosis in the neonatal rat spinal cord. J Neurol Sci 2001;185:109-118.
24.
Sen I, Nalini A, Joshi NB, Joshi PG: Cerebrospinal fluid from amyotrophic lateral sclerosis patients preferentially elevates intracellular calcium and toxicity in motor neurons via AMPA/kainate receptor. J Neurol Sci 2005;235:45-54.
25.
Shahani N, Gourie-Devi M, Nalini A, et al: (-)-Deprenyl alleviates the degenerative changes induced in the neonatal rat spinal cord by CSF from amyotrophic lateral sclerosis patients. Amyotroph Lateral Scler Other Motor Neuron Disord 2004;5:172-179.
26.
Anneser JM, Chahli C, Borasio GD: Protective effect of metabotropic glutamate receptor inhibition on amyotrophic lateral sclerosis-cerebrospinal fluid toxicity in vitro. Neuroscience 2006;141:1879-1886.
27.
Gunasekaran R, Narayani RS, Vijayalakshmi K, et al: Exposure to cerebrospinal fluid of sporadic amyotrophic lateral sclerosis patients alters Nav1.6 and Kv1.6 channel expression in rat spinal motor neurons. Brain Res 2009;1255:170-179.
28.
Orallo F: Trans-resveratrol: a magical elixir of eternal youth? Curr Med Chem 2008;15:1887-1898.
29.
Yáñez M, Galán L, Matias-Guiu J, Vela A, Guerrero A, García AG: CSF from amyotrophic lateral sclerosis patients produces glutamate independent death of rat motor brain cortical neurons: protection by resveratrol but not riluzole. Brain Res 2011;1423:77-86.
30.
Meininger V: European ALS Consortium. Amyotroph Lateral Scler Other Motor Neuron Disord 2004;5(suppl 1):133-134.
31.
Lanka V, Cudkowicz M: Therapy development for ALS: lessons learned and path forward. Amyotroph Lateral Scler 2008;9:131-140.
32.
Rodriguez de Rivera FJ, Grande M, Garcia-Caballero J, et al: Development of a clinical pathway for the attention of patients with amyotrophic lateral sclerosis in a regional network. ALS Assistance Network-Comunidad de Madrid (in Spanish). Neurologia 2007;22:354-361.
33.
Mosmann T: Rapid colorimetric assay for cellular growth and survival: application to proliferation and cytotoxicity assays. J Immunol Methods 1983;65:55-63.
34.
Vijayalakshmi K, Alladi PA, Ghosh S, et al: Evidence of endoplasmic reticular stress in the spinal motor neurons exposed to CSF from sporadic amyotrophic lateral sclerosis patients. Neurobiol Dis 2011;41:695-705.
35.
Vijayalakshmi K, Alladi PA, Sathyaprabha TN, Subramaniam JR, Nalini A, Raju TR: Cerebrospinal fluid from sporadic amyotrophic lateral sclerosis patients induces degeneration of a cultured motor neuron cell line. Brain Res 2009;1263:122-133.
36.
Kulshreshtha D, Vijayalakshmi K, Alladi PA, Sathyaprabha TN, Nalini A, Raju TR: Vascular endothelial growth factor attenuates neurodegenerative changes in the NSC-34 motor neuron cell line induced by cerebrospinal fluid of sporadic amyotrophic lateral sclerosis patients. Neurodegener Dis 2011;8:322-330.
37.
Hof PR, Perl DP: Neurofibrillary tangles in the primary motor cortex in Guamanian amyotrophic lateral sclerosis/parkinsonism-dementia complex. Neurosci Lett 2002;328:294-298.
38.
Fernandez-Santiago R, Hoenig S, Lichtner P, et al: Identification of novel angiogenin (ANG) gene missense variants in German patients with amyotrophic lateral sclerosis. J Neurol 2009;256:1337-1342.
39.
Shobha K, Vijayalakshmi K, Alladi PA, Nalini A, Sathyaprabha TN, Raju TR: Altered in-vitro and in-vivo expression of glial glutamate transporter-1 following exposure to cerebrospinal fluid of amyotrophic lateral sclerosis patients. J Neurol Sci 2007;254:9-16.
40.
Ramamohan PY, Gourie-Devi M, Nalini A, et al: Cerebrospinal fluid from amyotrophic lateral sclerosis patients causes fragmentation of the Golgi apparatus in the neonatal rat spinal cord. Amyotroph Lateral Scler 2007;8:79-82.
41.
Sankaranarayani R, Nalini A, Rao Laxmi T, Raju TR: Altered neuronal activities in the motor cortex with impaired motor performance in adult rats observed after infusion of cerebrospinal fluid from amyotrophic lateral sclerosis patients. Behav Brain Res 2010;206:109-119.
42.
Chen-Roetling J, Chen L, Regan RF: Minocycline attenuates iron neurotoxicity in cortical cell cultures. Biochem Biophys Res Commun 2009;386:322-326.
43.
Jorda EG, Verdaguer E, Canudas AM, et al: Implication of cyclin-dependent kinase 5 in the neuroprotective properties of lithium. Neuroscience 2005;134:1001-1011.
44.
Volbracht C, van Beek J, Zhu C, Blomgren K, Leist M: Neuroprotective properties of memantine in different in vitro and in vivo models of excitotoxicity. Eur J Neurosci 2006;23:2611-2622.
45.
Peluffo H, Estevez A, Barbeito L, Stutzmann JM: Riluzole promotes survival of rat motoneurons in vitro by stimulating trophic activity produced by spinal astrocyte monolayers. Neurosci Lett 1997;228:207-211.
46.
Arce MP, Rodriguez-Franco MI, Gonzalez-Munoz GC, et al: Neuroprotective and cholinergic properties of multifunctional glutamic acid derivatives for the treatment of Alzheimer's disease. J Med Chem 2009;52:7249-7257.
47.
Arias E, Ales E, Gabilan NH, et al: Galantamine prevents apoptosis induced by beta-amyloid and thapsigargin: involvement of nicotinic acetylcholine receptors. Neuropharmacology 2004;46:103-114.
48.
Arias E, Gallego-Sandin S, Villarroya M, Garcia AG, Lopez MG: Unequal neuroprotection afforded by the acetylcholinesterase inhibitors galantamine, donepezil, and rivastigmine in SH-SY5Y neuroblastoma cells: role of nicotinic receptors. J Pharmacol Exp Ther 2005;315:1346-1353.
49.
Bond M, Rogers G, Peters J, et al: The effectiveness and cost-effectiveness of donepezil, galantamine, rivastigmine and memantine for the treatment of Alzheimer's disease (review of Technology Appraisal No 111): a systematic review and economic model. Health Technol Assess 2012;16:1-470.
50.
Wang JL, Lee KC, Tang KY, et al: Effect of the neuroprotective agent riluzole on intracellular Ca2+ levels in IMR32 neuroblastoma cells. Arch Toxicol 2001;75:214-220.
51.
Jan CR, Lu YC, Jiann BP, Chang HT, Huang JK: Effect of riluzole on cytosolic Ca2+ increase in human osteosarcoma cells. Pharmacology 2002;66:120-127.
52.
Chen WC, Cheng HH, Huang CJ, et al: Effect of riluzole on Ca2+ movement and cytotoxicity in Madin-Darby canine kidney cells. Hum Exp Toxicol 2006;25:461-469.
53.
Akamatsu K, Shibata MA, Ito Y, Sohma Y, Azuma H, Otsuki Y: Riluzole induces apoptotic cell death in human prostate cancer cells via endoplasmic reticulum stress. Anticancer Res 2009;29:2195-2204.
54.
Caumont AS, Octave JN, Hermans E: Specific regulation of rat glial cell line-derived neurotrophic factor gene expression by riluzole in C6 glioma cells. J Neurochem 2006;97:128-139.
55.
Mizuta I, Ohta M, Ohta K, Nishimura M, Mizuta E, Kuno S: Riluzole stimulates nerve growth factor, brain-derived neurotrophic factor and glial cell line-derived neurotrophic factor synthesis in cultured mouse astrocytes. Neurosci Lett 2001;310:117-120.
56.
Katoh-Semba R, Takeuchi IK, Inaguma Y, et al: Induction of brain-derived neurotrophic factor by convulsant drugs in the rat brain: involvement of region-specific voltage-dependent calcium channels. J Neurochem 2001;77:71-83.
57.
Milane A, Tortolano L, Fernandez C, Bensimon G, Meininger V, Farinotti R: Brain and plasma riluzole pharmacokinetics: effect of minocycline combination. J Pharm Pharm Sci 2009;12:209-217.
58.
Bellingham MC: A review of the neural mechanisms of action and clinical efficiency of riluzole in treating amyotrophic lateral sclerosis: what have we learned in the last decade? CNS Neurosci Ther 2010;17:4-31.
59.
Meininger V, Bensimon G, Bradley WR, et al: Efficacy and safety of xaliproden in amyotrophic lateral sclerosis: results of two phase III trials. Amyotroph Lateral Scler Other Motor Neuron Disord 2004;5:107-117.
60.
Rowland LP: Riluzole for the treatment of amyotrophic lateral sclerosis - too soon to tell? N Engl J Med 1994;330:636-637.
61.
Rowland LP, Shneider NA: Amyotrophic lateral sclerosis. N Engl J Med 2001;344:1688-1700.
62.
Meininger V: Clinical trials in ALS: what did we learn from recent trials in humans? Neurodegener Dis 2005;2:208-214.
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2013
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