Biomarkers in amyotrophic lateral sclerosis: combining metabolomic and clinical parameters to define disease progression.

BACKGROUND AND PURPOSE: The objectives of this study were to define the metabolomic profile of cerebrospinal fluid in amyotrophic lateral sclerosis (ALS) patients, to model outcome through combined clinical and metabolomic parameters and independently to validate predictive models. METHODS: In all, 74 consecutive newly diagnosed patients were enrolled into training (Tr, n = 49) and test (Te, n = 25) cohorts. Investigators recorded clinical data and the metabalomic profile of cerebrospinal fluid at baseline was analyzed with (1)H nuclear magnetic resonance spectroscopy. Markers of disease progression, collected in 1-year prospective follow-up, included change in ALS Functional Rating Scale (var_ALSFRS), change in weight (var_weight) and survival time. Stepwise multiple regression selected from metabolomic and clinical parameters to model rate of progression in the Tr cohort. Best fit models were validated independently in the Te cohort. RESULTS: The best-fit statistical models, using both metabolomic and clinical covariates, predicted outcome with 70.8% (var_weight), 72% (var_ALSFRS) and 76% (survival) accuracy in the Te cohort. Models that used metabolomics or clinical data alone predicted outcome less well. Highlighted metabolites are involved in pathophysiological pathways previously described in ALS. CONCLUSION: Cerebrospinal fluid metabolomics can aid in predicting the clinical course of ALS and tap into pathophysiological processes. The precision of predictive models, independently reproduced in this study, is enhanced through inclusion of both metabolomic and clinical parameters. The findings bring the field closer to a clinically meaningful disease marker.

The glutamate hypothesis in ALS: pathophysiology and drug development.

Amyotrophic lateral sclerosis (ALS) is an age-related neurodegenerative disorder that is believed to have complex genetic and environmental influences in the pathogenesis, but etiologies are unidentified for most patients. Until the major causes are better defined, drug development is directed at downstream pathophysiological mechanisms, themselves incompletely understood. For nearly 30 years, glutamate-induced excitotoxicity has lain at the core of theories behind the spiraling events, including mitochondrial dysfunction, oxidative stress, and protein aggregation, that lead to neurodegenerative cell death. One drug, riluzole, which possesses anti-glutamatergic properties, is approved as neuroprotective for ALS. Following the achievement of the riluzole trials, numerous other agents with similar mechanisms have been tested without success. This article provides an overview of excitotoxicity in ALS, focusing on the events that contribute to excess glutamate, how the excess might damage nerve cells, and how this information is being harnessed in the development of potential new neuroprotective agents. The work highlights clinical trials of drugs that have targeted the glutamate system, comments on the potential role of glutamate as a biomarker and concludes with a section on future directions for the field. As research uncovers elusive etiologies and brings clarity to pathophysiological mechanisms, the success of new interventions will increasingly depend on the design of agents that target particular mechanisms for specific individuals. The heady future of personalized drug regimens for ALS rests with medicinal chemists, the scientists whose ideas and work produce these designer drugs.

Synthesis of imidazo[1,2-a]pyridine derivatives as antiviral agents.

The synthesis and antiviral activity of original dibromoimidazo[1,2-a]pyridines bearing a thioether side chain are reported. Molecular modeling was used to identify biophoric structural patterns that are common to 16 compounds. Structure-activity relationship (SAR) studies identified hydrophobicity (logP) as the most important factor for activity. From these SAR studies, the antiviral activity could be predicted.

Synthesis and cytotoxicity of novel pyrido[1,2-e]purines on multidrug resistant human MCF7 cells.

The anticancer activity of 4-methylaminopyridol[1,2-e]purine 6a, 4-(piperidin-1-yl)pyrido[1,2-e]purine 7a and their 7-methyl derivatives 6b, 7b was investigated against the human MCF7 cancer cell line in vitro. The sensitive cell line showed a range of sensitivities to 6a, 6b, 7a, 7b (IC50: 1.6 to 7.2 x 10(-4) M) and sensitivity to doxorubicin (IC50: 7.5 x 10(-7) M). A resistant cell line with the multidrug resistant phenotype was sensitive to these derivatives (IC50: 1.8 to 6.7 x 10(-4) M), doxorubicin (IC50: 5 x 10(-5) M) and drug activity seems to be not affected by MDR resistance. Our data show that 6a, 6b, 7a and 7b appear to exert a low cytotoxicity on sensitive and MDR resistant MCF7 human cancer cell lines.

Synthesis of imidazo[1,2-a]pyridines as antiviral agents.

The synthesis of original imidazo[1,2-a]pyridines bearing a thioether side chain at the 3 position and their antiviral activity are reported. From the synthesized compounds, 4, 15, and 21 were highly active against human cytomegalovirus with a therapeutic index superior to 150. These compounds also showed pronounced activity against varicella-zoster virus. Their structure-activity relationship is discussed.

Effects of two N-arylpiperazinylmethylpyrazolo [1,5-d][1,2,4]triazine derivatives in pain and antidepressant tests in mice.

The antinociceptive and antidepressant effects of two pyrazolotriazine derivatives, 2-phenyl-3,3a-dihydro-4-oxo-5-(4-phenylpiperazin-1-yl) methyl-pyrazolo[1,5-d][1,2,4]-triazine (SM1) and 2-phenyl-3,3a-dihydro-4-oxo-5-[4-(4-fluorophenyl)piperazin-1-yl] methylpyrazolo[1,5-d][1,2,4] triazine (SM3) have been investigated in mice using classical pharmacological tests. The intraperitoneal LD50 values of SM1 and SM3 were 253.4 and 218.8 mg kg-1 respectively. SM1 and SM3 showed analgesic properties in the phenylbenzoquinone-induced abdominal constriction test (ED50 approximately 10-15 mg kg-1, i.p.) and in the hot-plate test. The antinociceptive effects of the triazines were significantly reduced by administration of naloxone (1 and 3.2 mg kg-1, s.c.) and yohimbine (1 mg kg-1, p.o.). Acute intraperitoneal administration of both compounds (1 mg kg-1 SM1 or 1.5 mg kg-1 SM3) potentiated morphine (0.15 mg kg-1, s.c.) analgesia in the phenylbenzoquinone test. Although this synergistic activity was not reversed by methysergide (0.5 mg kg-1, i.p.), the analgesic activity of both compounds was enhanced by administration of 5-hydroxytryptophan (50 mg kg-1, i.p.) in conjunction with carbidopa (25 mg kg-1, i.p.). Furthermore, neither compound (at 100 mg kg-1, i.p.) significantly reduced the duration of immobility of mice in the forced swimming test, and both (at 75 mg kg-1, i.p.) were ineffective at enhancing the toxic effects of yohimbine (30 mg kg-1, s.c.). Only SM3 (ED50 = 74.5 mg kg-1, i.p.) significantly antagonized reserpine (2.5 mg kg-1, i.p.)-induced ptosis. Thus, the results suggest that SM1 and SM3 have antinociceptive properties related to co-involvement of opioidergic and alpha 2-adrenoceptor mechanism without associated antidepressant properties.

Synthesis and pharmacological evaluation in mice of new non-classical antinociceptive agents, 5-(4-arylpiperazin-1-yl)-4-benzyl-1,2-oxazin-6-ones.

Several 5-(4-arylpiperazin-1-yl)-4-benzyl-1,2-oxazin-6-ones have been synthesized and tested for analgesic activity in a visceral pain model (phenylbenzoquinone-induced writhing test = PBQ test). A good correlation has been found between the antinociceptive effects of drugs and both their lipophilic and steric properties. The most active derivatives 5c and 5f, with intraperitoneal ED50 values of 10.5 and 10.3 mg kg-1 respectively, were more extensively investigated by evaluating their analgesic activity in a somatosensory pain model (hot plate test), as well as their sedative properties. Furthermore, naloxone suppressed the effect of 5c and 5f in the PBQ test, though these derivatives were ineffective to potentiate morphine analgesia. Pretreatment with yohimbine did not significantly attenuate the analgesic effects of 5c and 5f. In addition, pretreatment with 5-hydroxytryptophan associated with carbidopa also failed to potentiate the antinociceptive effects of 5c and 5f. So, a part of the analgesic activity of 5c and 5f seems to be related to an opioidergic mechanisms, especially at the mu receptor level. Molecular modeling studies performed on the opiate drug morphine and on the most stable conformer of 5f showed structural similarities between these two molecules.

Thromboxane A2 biosynthesis inhibitors: synthesis and evaluation of pyrazolotriazinyl alkanoic acids.

A novel series of (6-aryl-4-oxo-pyrazolo 2,3-d] [1,2,5] triazin-3-yl) alkanoic acids was synthesized and evaluated in vitro as thromboxane A2 (TXA2) biosynthesis inhibitors. The experiments were carried out using arachidonic acid (32.8 microM) as a substrate and horse platelet microsomes (HPM) as sources of TXA synthetase. TXB2, a stable breakdown product of TXA2, was determined by radioimmunoassays (RIA). The substances under study, at concentrations ranging from 1.10(-6) M to 1.10(-4) M, significantly inhibited the biosynthesis of TXA2 in vitro. This activity was found to be dose-dependent, the potency of which could be related to structural features of the molecules. Compound 3b, bearing a butanoic side chain in the 3-position and a 4-chloro phenyl ring in the 6-position of the bicyclic system, was the most active derivative in in vitro enzyme inhibition (ID50 = 2.81 x 10(-5) M). Comparison of the spatial configurations of prostaglandin H2 (PGH2 and 3b displayed a good correlation between essential structural moieties of both molecules. In addition, conceptual model for the PGH2 and TX synthetase interactions was applied to compound 3b.

Synthesis of new 5-substituted pyrazolo[1,5-d][1,2,4]-triazine derivatives and their analgesic, anti-inflammatory and antipyretic properties.

A series of 2-aryl-4-oxo-pyrazolo[1,5-d][1,2,4]triazines substituted in the 5-position by aminoalkyl or benzoyl moieties was synthesized and evaluated for analgesic activity. The structures of new triazine derivatives were confirmed by IR, 1H-NMR spectra and by elementary analysis. In the phenylbenzoquinone induced writhing test, only 3,3a-dihydropyrazolo triazines substituted by an arylpiperazinylmethyl group exhibited potent analgesic effect. In addition, these compounds possessed significant anti-inflammatory and antipyretic properties. A desaturation in 3,3a positions or other groups than arylpiperazinylmethyl moieties notably decreased analgesic effects.

Synthesis and aldose reductase inhibitory activity of triazine derivatives possessing acetic acid group.

N-Acetic acid derivatives of 6-aryl-pyrazolo-triazin-4-ones were synthesized for evaluation as new aldose reductase inhibitors. The intrinsic activity of each compound was assessed by measuring the inhibition of enzymatic activity in an isolated pig lens enzyme preparation. All the prepared compounds exhibited a significant in vitro aldose reductase inhibitory effect (10(-6) M less than or equal to IC50 less than or equal to 10(-4) M). Furthermore, biological activity (log 1/IC50) for most of the data sets could be correlated directly to electronic and steric parameters. Finally, spatial configuration of the most active derivative 6c (IC50 = 2 x 10(-6) M) was compared with that of tolrestat and with pharmacophor requirements of the aldose reductase inhibitor site using a molecular modeling system.