Can Some Anticancer Drugs Be Repurposed to Treat Amyotrophic Lateral Sclerosis? A Brief Narrative Review.

Amyotrophic lateral sclerosis (ALS) is a rare progressive motor neuron disease that, due to its high complexity, still lacks effective treatments. Development of a new drug is a highly costly and time-consuming process, and the repositioning of approved drugs can represent an efficient strategy to provide therapeutic opportunities. This is particularly true for rare diseases, which are characterised by small patient populations and therefore attract little commercial interest. Based on the overlap between the biological background of cancer and neurodegeneration, the repurposing of antineoplastic drugs for ALS has been suggested. The objective of this narrative review was to summarise the current experimental evidence on the use of approved anticancer drugs in ALS. Specifically, anticancer drugs belonging to different classes were found to act on mechanisms involved in the ALS pathogenesis, and some of them proved to exert beneficial effects in ALS models. However, additional studies are necessary to confirm the real therapeutic potential of anticancer drugs for repositioning in ALS treatment.

Fenretinide Beneficial Effects on Amyotrophic Lateral Sclerosis-associated SOD1G93A Mutant Protein Toxicity: In Vitro and In Vivo Evidences.

Amyotrophic lateral sclerosis (ALS) is the most frequent motor neuron disease for which effective treatment options are still lacking. ALS occurs in sporadic and familial forms which are clinically indistinguishable; about 20% of familial ALS cases are linked to mutations of the superoxide dismutase 1 (SOD1) gene. Fenretinide (FEN), a cancer chemopreventive and antiproliferative agent currently used in several clinical trials, is a multi-target drug which also exhibits redox regulation activities. We analyzed the effects of FEN on mutant SOD1 (mSOD1) toxicity in motoneuronal (NSC34) and a muscle (C2C12) cell lines and evaluated the impacts of chronic administration of a new nanomicellar fenretinide formulation (NanoMFen) on ALS disease progression in the SOD1G93A mouse model. The results showed that FEN significantly prevents the toxicity of mSOD1 expression in NSC34 motor neuron; furthermore, FEN is able to partially overcome the toxic effect of mSOD1 on the myogenic program of C2C12 muscle cells. Administration of NanoMFen ameliorates the disease progression and increases median survival of mSOD1G93A ALS mice, even when given after disease onset; beneficial effects in ALS mice, however, is restricted to female sex. Our data support the therapeutic potential of FEN against ALS-associated SOD1G93A mutant protein toxicity and promote further studies to elucidate specific cellular targets of the drug in ALS. Furthermore, the sex-related efficacy of NanoMFen in mSOD1G93A ALS mice strengthens the importance, in the perspective of a precision medicine approach, of gender pharmacology in ALS research.

Adenosine A2A receptor inhibition reduces synaptic and cognitive hippocampal alterations in Fmr1 KO mice.

In fragile X syndrome (FXS) the lack of the fragile X mental retardation protein (FMRP) leads to exacerbated signaling through the metabotropic glutamate receptors 5 (mGlu5Rs). The adenosine A2A receptors (A2ARs), modulators of neuronal damage, could play a role in FXS. A synaptic colocalization and a strong permissive interaction between A2A and mGlu5 receptors in the hippocampus have been previously reported, suggesting that blocking A2ARs might normalize the mGlu5R-mediated effects of FXS. To study the cross-talk between A2A and mGlu5 receptors in the absence of FMRP, we performed extracellular electrophysiology experiments in hippocampal slices of Fmr1 KO mouse. The depression of field excitatory postsynaptic potential (fEPSPs) slope induced by the mGlu5R agonist CHPG was completely blocked by the A2AR antagonist ZM241385 and strongly potentiated by the A2AR agonist CGS21680, suggesting that the functional synergistic coupling between the two receptors could be increased in FXS. To verify if chronic A2AR blockade could reverse the FXS phenotypes, we treated the Fmr1 KO mice with istradefylline, an A2AR antagonist. We found that hippocampal DHPG-induced long-term depression (LTD), which is abnormally increased in FXS mice, was restored to the WT level. Furthermore, istradefylline corrected aberrant dendritic spine density, specific behavioral alterations, and overactive mTOR, TrkB, and STEP signaling in Fmr1 KO mice. Finally, we identified A2AR mRNA as a target of FMRP. Our results show that the pharmacological blockade of A2ARs partially restores some of the phenotypes of Fmr1 KO mice, both by reducing mGlu5R functioning and by acting on other A2AR-related downstream targets.

Modulating P1 Adenosine Receptors in Disease Progression of SOD1G93A Mutant Mice.

Amyotrophic lateral sclerosis (ALS) is a fatal progressing neurodegenerative disease; to date, despite the intense research effort, only two therapeutic options, with very limited effects, are available. The purinergic system has been indicated as a possible new therapeutic target for ALS, but the results are often contradictory and generally confused. The present study was designed to determine whether P1 adenosine receptor ligands affected disease progression in a transgenic model of ALS. SOD1G93A mice were chronically treated, from presymptomatic stage, with a selective adenosine A2A receptor agonist (CGS21680), antagonist (KW6002) or the A1 receptor antagonist DPCPX. Body weight, motor performance and survival time were evaluated. The results showed that neither the stimulation nor the blockade of adenosine A2A receptors modified the progressive loss of motor skills or survival of mSOD1G93A mice. Conversely, blockade of adenosine A1 receptors from the presymptomatic stage significantly attenuated motor disease progression and induced a non-significant increase of median survival in ALS mice. Our data confirm that the modulation of adenosine receptors can elicit very different (and even opposite) effects during the progression of ALS course, thus strengthens the importance of further studies to elucidated their real therapeutic potential in this pathology.

Neuronal adenosine A2A receptor overexpression is neuroprotective towards 3-nitropropionic acid-induced striatal toxicity: a rat model of Huntington's disease.

The A2A adenosine receptor (A2AR) is widely distributed on different cellular types in the brain, where it exerts a broad spectrum of pathophysiological functions, and for which a role in different neurodegenerative diseases has been hypothesized or demonstrated. To investigate the role of neuronal A2ARs in neurodegeneration, we evaluated in vitro and in vivo the effect of the neurotoxin 3-nitropropionic acid (3-NP) in a transgenic rat strain overexpressing A2ARs under the control of the neural-specific enolase promoter (NSEA2A rats). We recorded extracellular field potentials (FP) in corticostriatal slice and found that the synaptotoxic effect of 3-NP was significantly reduced in NSEA2A rats compared with wild-type animals (WT). In addition, after exposing corticostriatal slices to 3-NP 10 mM for 2 h, we found that striatal cell viability was significantly higher in NSEA2A rats compared to control rats. These in vitro results were confirmed by in vivo experiments: daily treatment of female rats with 3-NP 10 mg/kg for 8 days induced a selective bilateral lesion in the striatum, which was significantly reduced in NSEA2A compared to WT rats. These results demonstrate that the overexpression of the A2AR selectively at the neuronal level reduced 3-NP-induced neurodegeneration, and suggest an important function of the neuronal A2AR in the modulation of neurodegeneration.

The adenosine A2A receptor agonist T1-11 ameliorates neurovisceral symptoms and extends the lifespan of a mouse model of Niemann-Pick type C disease.

Niemann-Pick C is a fatal neurovisceral disorder caused, in 95% of cases, by mutation of NPC1 gene. Therapeutic options are extremely limited and new "druggable" targets are highly warranted. We previously demonstrated that the stimulation of the adenosine A2A receptor (A2AR) normalized the pathological phenotype of cellular models of NPC1. Since the validation of A2ARs as a therapeutic target for NPC1 can be obtained only conducting studies in in vivo models of the disease, in the present paper, the effects of two agonists of A2ARs were evaluated in the mouse model Balb/c Npc1nih, hereafter indicated as NPC1-/-. The agonists CGS21680 (2.5 and 5mg/kg/day by intraperitoneal injection) and T1-11 (50mg/kg/day in drinking water) were administered at a presymptomatic stage of the disease of NPC1-/- mice (PN28 and PN30, respectively); the experimental groups were the following: vehicle-treated WT mice (N=16 for both CGS and T1-11 treatments); vehicle-treated NPC1-/- mice (N=14 for CGS and 12 for T1-11 treatment); CGS-treated NPC1-/- mice (N=7) and T1-11-treated NPC1-/- mice (N=11). The efficacy of the treatments was evaluated by comparing vehicle-treated and CGS or T1-11-treated NPC1-/- mice for their motor deficits (analyzed by both rotarod and footprint tests), hippocampal cognitive impairment (by Novel Object Recognition (NOR) test), cerebellar neurodegeneration (Purkinje neurons counting), and cholesterol and sphingomyelin accumulation in spleen and liver. Finally, the effect of both agonists on survival was evaluated by applying a humane late endpoint (weight loss >30% of peak weight, punched posture and reduced activity in the cage). The results demonstrated that, while CGS21680 only slightly attenuated cognitive deficits, T1-11 ameliorated motor coordination, significantly improved cognitive impairments, increased the survival of Purkinje neurons and reduced sphingomyelin accumulation in the liver. More importantly, it significantly prolonged the lifespan of NPC1-/- mice. In vitro experiments conducted in a neuronal model of NPC1 demonstrated that the ability of T1-11 to normalize cell phenotype was mediated by the selective activation of A2ARs and modulation of intracellular calcium levels. In conclusion, our results fully confirm the validity of A2ARs as a new target for NPC1 treatment. As soon as new ligands with improved pharmacokinetic characteristics (i.e. orally active, with brain bioavailability and metabolic stability) will be obtained, A2AR agonists could represent a breakthrough in the treatment of NPC.

Striatal adenosine-cannabinoid receptor interactions in rats over-expressing adenosine A2A receptors.

Adenosine A2A receptors (A2 A Rs) and cannabinoid CB1 receptors (CB1 Rs) are highly expressed in the striatum, where they functionally interact and form A2A /CB1 heteroreceptor complexes. We investigated the effects of CB1 R stimulation in a transgenic rat strain over-expressing A2 A Rs under the control of the neural-specific enolase promoter (NSEA2A rats) and in age-matched wild-type (WT) animals. The effects of the CB1 R agonist WIN 55,212-2 (WIN) were significantly lower in NSEA2A rats than in WT animals, as demonstrated by i) electrophysiological recordings of synaptic transmission in corticostriatal slices; ii) the measurement of glutamate outflow from striatal synaptosomes and iii) in vivo experiments on locomotor activity. Moreover, while the effects of WIN were modulated by both A2 A R agonist (CGS 21680) and antagonists (ZM 241385, KW-6002 and SCH-442416) in WT animals, the A2 A R antagonists failed to influence WIN-mediated effects in NSEA2A rats. The present results demonstrate that in rats with genetic neuronal over-expression of A2 A Rs, the effects mediated by CB1 R activation in the striatum are significantly reduced, suggesting a change in the stoichiometry of A2A and CB1 receptors and providing a strategy to dissect the involvement of A2 A R forming or not forming heteromers in the modulation of striatal functions. These findings add additional evidence for the existence of an interaction between striatal A2 A Rs and CB1 Rs, playing a fundamental role in the regulation of striatal functions. We studied A2A -CB1 receptor interaction in transgenic rats over-expressing adenosine A2A receptors under the control of the neuron-specific enolase promoter (NSEA2A ). In these rats, we demonstrated a reduced effect of the CB1 receptor agonist WIN 55,212-2 in the modulation of corticostriatal synaptic transmission and locomotor activity, while CB1 receptor expression level did not change with respect to WT rats. A reduction in the expression of A2A -CB1 receptor heteromers is postulated.

Effects of chronic caffeine intake in a mouse model of amyotrophic lateral sclerosis.

Caffeine is a nonselective adenosine receptor antagonist; chronic consumption has proved protective toward neurodegenerative diseases such as Parkinson's and Alzheimer's diseases. The present study was designed to determine whether caffeine intake affected survival and/or motor performance in a transgenic model of amyotrophic lateral sclerosis (ALS). SOD1(G93A) mice received caffeine through drinking water from 70 days of age until death. Body weight, motor performance and survival were evaluated. Furthermore, the expression of adenosine A(2A) receptors (A(2A) Rs), glial glutamate transporter (GLT1), and glial fibrillar acidic protein (GFAP) were evaluated by Western blotting. The results showed that caffeine intake significantly shortened the survival of SOD1(G93A) mice (log rank test, P = 0.01) and induced a nonsignificant advancing of disease onset. The expression of A(2A) R, GLT1, and GFAP was altered in the spinal cords of ALS mice, but caffeine did not influence their expression in either wild-type or SOD1(G93) mice. These data indicate that adenosine receptors may play an important role in ALS.

Influence of CGS 21680, a selective adenosine A(2A) receptor agonist, on NMDA receptor function and expression in the brain of Huntington's disease mice.

The effect of chronic treatment with the selective adenosine A(2A) receptor agonist CGS 21680 on N-Methyl-d-Aspartate (NMDA) receptor function and expression has been studied in the striatum and cortex of R6/2 mice, a genetic mouse model of Huntington's disease (HD). Starting from 8weeks of age, R6/2 and wild type (WT) mice were treated daily with CGS 21680 (0.5mg/kg i.p.) for 3weeks and the expression levels of NMDA receptor subunits were then evaluated. In addition, to study CGS 21680-induced changes in NMDA receptor function, NMDA-induced toxicity in corticostriatal slices from both R6/2 and WT mice was investigated. We found that CGS 21680 increased NR2A subunit expression and the NR2A/NR2B ratio in the cortex of R6/2 mice, having no effect in WT mice. In the striatum, CGS 21680 reduced NR1 expression in both R6/2 and WT mice while the effect on NR2A and NR2/NR2B expression was genotype-dependent, reducing and increasing their expression in WT and R6/2 mice, respectively. On the contrary, NMDA-induced toxicity in corticostriatal slices was not modified by the treatment in WT or HD mice. These results demonstrate that in vivo activation of A(2A) receptors modulates the subunit composition of NMDA receptors in the brain of HD mice.

Remodeling of striatal NMDA receptors by chronic A(2A) receptor blockade in Huntington's disease mice.

Excitotoxicity plays a major role in the pathogenesis of Huntington disease (HD), a fatal neurodegenerative disorder. Adenosine A(2A) receptors (A(2A)Rs) modulate excitotoxicity and have been suggested to play a pathogenetic role in HD. The main aim of this study was to evaluate the effect of A(2A)R blockade on the expression and functions of NMDA receptors in the striatum of HD mice (R6/2). We found that 3 weeks' treatment with SCH 58261 (0.01 mg/kg/day i.p. from the 8th week of age) modified NR1 and NR2A/NR2B expression in the striatum of R6/2 (Western blotting) while had no effect on NMDA-induced toxicity in corticostriatal slices (electrophysiological experiments). In conclusion, in vivo A(2A)R blockade induced a remodeling of NMDA receptors in the striatum of HD mice. Even though the functional relevance of the above effect remains to be fully elucidated, these results add further evidence to the modulatory role of A(2A)Rs in HD.