Neuroprotective and neurorestorative potential of propargylamine derivatives in ageing: focus on mitochondrial targets.

The mitochondrial theory of ageing proposes that accumulation of damage to mitochondrial function and DNA mutation lead to ageing of humans and animals. It has been suggested that mitochondria play dynamic roles in regulating synaptogenesis and morphological/functional responses of synaptic activity, and thus, deteriorating of mitochondrial function (e.g., deficits of the mitochondrial respiratory enzymes, reduced calcium influx, increased accumulation of mitochondrial DNA defects/apoptotic proteins and impairment of mitochondrial membrane potential) can lead to severe neuronal energy deficit, and in the long run, to modifications in neuronal synapses and neurodegeneration in the ageing brain. Hence, considering the mechanisms by which mitochondrial impairment can lead to neuronal death, the development of neuroprotective molecules that target various mitochondrial pathogenic processes can be effective in the treatment of ageing and age-related neurodegenerative diseases. This review addresses several aspects of the neuroprotective effects of propargylamine derivatives (e.g., the monoamine oxidase-B inhibitors, selegiline and rasagiline and the multifunctional drugs, ladostigil, M30 and VAR10303) in ageing with a special focus on mitochondrial molecular protective mechanisms.

Development of a local controlled release system for therapeutic proteins in the treatment of skeletal muscle injuries and diseases.

The present study aims to develop and characterize a controlled-release delivery system for protein therapeutics in skeletal muscle regeneration following an acute injury. The therapeutic protein, a membrane-GPI anchored protein called Cripto, was immobilized in an injectable hydrogel delivery vehicle for local administration and sustained release. The hydrogel was made of poly(ethylene glycol)-fibrinogen (PEG-Fibrinogen, PF), in the form of injectable microspheres. The PF microspheres exhibited a spherical morphology with an average diameter of approximately 100 micrometers, and the Cripto protein was uniformly entrapped within them. The release rate of Cripto from the PF microspheres was controlled by tuning the crosslinking density of the hydrogel, which was varied by changing the concentration of poly(ethylene glycol) diacrylate (PEG-DA) crosslinker. In vitro experiments confirmed a sustained-release profile of Cripto from the PF microspheres for up to 27 days. The released Cripto was biologically active and promoted the in vitro proliferation of mouse myoblasts. The therapeutic effect of PF-mediated delivery of Cripto in vivo was tested in a cardiotoxin (CTX)-induced muscle injury model in mice. The Cripto caused an increase in the in vivo expression of the myogenic markers Pax7, the differentiation makers eMHC and Desmin, higher numbers of centro-nucleated myofibers and greater areas of regenerated muscle tissue. Collectively, these results establish the PF microspheres as a potential delivery system for the localized, sustained release of therapeutic proteins toward the accelerated repair of damaged muscle tissue following acute injuries.

Effects of novel neuroprotective and neurorestorative multifunctional drugs on iron chelation and glucose metabolism.

Iron accumulation and iron-related oxidative stress are involved in several pathological conditions and provide a rationale for the development of iron chelators as novel promising therapeutic strategies. Thus, we have recently synthesized multifunctional non-toxic, brain permeable iron chelating compounds, M30 and HLA20, possessing the neuroprotective N-propargyl moiety of the anti-Parkinsonian drug, monoamine oxidase (MAO)-B inhibitor, rasagiline and the antioxidant-iron chelating moiety of an 8-hydroxyquinoline derivative of the iron chelator, VK28. Here, we examined the hepatic regulatory effects of these novel compounds using two experimental approaches: chelation activity and glucose metabolism parameters. The present study demonstrated that M30 and HLA20 significantly decreased intracellular iron content and reduced ferritin expression levels in iron-loaded hepatoma Hep3B cells. In electron microscopy analysis, M30 was shown to reduce the electron-dense deposits of siderosomes by ~30 %, as well as down-regulate cytosolic ferritin particles observed in iron-overloaded cells. In vivo studies demonstrated that M30 administration (1 mg/kg, P.O. three times a week) reduced hepatic ferritin levels; increased hepatic insulin receptor and glucose transporter-1 levels and improved glucose tolerance in C57BL/6 mice and in a mouse model of type-2 diabetes, the ob/ob (leptin(-/-)). The results clearly indicate that the novel multifunctional drugs, especially M30, display significant capacity of chelating intracellular iron and regulating glucose metabolism parameters. Such effects can have therapeutic significance in conditions with abnormal local or systemic iron metabolism, including neurological diseases.

Methacrylated fibrinogen hydrogels for 3D cell culture and delivery.

Methacrylation was performed on fibrinogen to design a new biomedical hydrogel for 3D cell culture or as a biodegradable delivery matrix for in vivo implantation. The methacrylation of denatured fibrinogen in solution was performed using methacrylic anhydride (MAA). The extent of fibrinogen methacrylation was quantified by proton NMR and controlled using stochiometric quantities of MAA during the reaction. The methacrylated fibrinogen (FibMA) hydrogels were formed by light-activated free-radical polymerization in the presence of macromolecular cross-linking polymers made from acrylated poly(ethylene glycol) (PEG). The biocompatibility and biodegradability of the FibMA hydrogels were characterized by in vitro assays and in vivo implantation experiments using quantitative magnetic resonance imaging (MRI) of the implant volume. The FibMA supported the growth and metabolic activity of human dermal fibroblasts in both 2D and 3D cultures. The methacrylation did not alter important biological attributes of the fibrinogen, including the ability to support cell adhesion and 3D cell culture, as well as to undergo proteolysis. Animal experiments confirmed the biodegradability of the FibMA for potential use as a scaffold in tissue engineering, as a bioink for 3D bioprinting, or as a biodegradable matrix for in vivo sustained delivery of bioactive factors. STATEMENT OF SIGNIFICANCE: This paper describes methacrylated fibrinogen (FibMA) and the formation of a biomedical hydrogel from FibMA for cell culture and other biomedical applications. Inspired from methacrylated gelatin (GelMA), the FibMA is made from blood-derived fibrinogen which is more suitable for clinical use. Sharing similar properties to other hydrogels made from methacrylated proteins, the FibMA has yet to be reported in the literature. In this manuscript, we provide the methodology to produce the FibMA hydrogels, we document the mechanical versatility of this new biomaterial, and we show the biocompatibility using 3D cell culture studies and in vivo implantations.

Biomanufacturing Recombinantly Expressed Cripto-1 Protein in Anchorage-Dependent Mammalian Cells Growing in Suspension Bioreactors within a Three-Dimensional Hydrogel Microcarrier.

Biotherapeutic soluble proteins that are recombinantly expressed in mammalian cells can pose a challenge when biomanufacturing in three-dimensional (3D) suspension culture systems. Herein, we tested a 3D hydrogel microcarrier for a suspension culture of HEK293 cells overexpressing recombinant Cripto-1 protein. Cripto-1 is an extracellular protein that is involved in developmental processes and has recently been reported to have therapeutic effects in alleviating muscle injury and diseases by regulating muscle regeneration through satellite cell progression toward the myogenic lineage. Cripto-overexpressing HEK293 cell lines were cultured in microcarriers made from poly (ethylene glycol)-fibrinogen (PF) hydrogels, which provided the 3D substrate for cell growth and protein production in stirred bioreactors. The PF microcarriers were designed with sufficient strength to resist hydrodynamic deterioration and biodegradation associated with suspension culture in stirred bioreactors for up to 21 days. The yield of purified Cripto-1 obtained using the 3D PF microcarriers was significantly higher than that obtained with a two-dimensional (2D) culture system. The bioactivity of the 3D-produced Cripto-1 was equivalent to commercially available Cripto-1 in terms of an ELISA binding assay, a muscle cell proliferation assay, and a myogenic differentiation assay. Taken together, these data indicate that 3D microcarriers made from PF can be combined with mammalian cell expression systems to improve the biomanufacturing of protein-based therapeutics for muscle injuries.

In vivo restoration of dystrophin expression in mdx mice using intra-muscular and intra-arterial injections of hydrogel microsphere carriers of exon skipping antisense oligonucleotides.

Duchenne muscular dystrophy (DMD) is a genetic disease caused by a mutation in the X-linked Dytrophin gene preventing the expression of the functional protein. Exon skipping therapy using antisense oligonucleotides (AONs) is a promising therapeutic strategy for DMD. While benefits of AON therapy have been demonstrated, some challenges remain before this strategy can be applied more comprehensively to DMD patients. These include instability of AONs due to low nuclease resistance and poor tissue uptake. Delivery systems have been examined to improve the availability and stability of oligonucleotide drugs, including polymeric carriers. Previously, we showed the potential of a hydrogel-based polymeric carrier in the form of injectable PEG-fibrinogen (PF) microspheres for delivery of chemically modified 2'-O-methyl phosphorothioate (2OMePs) AONs. The PF microspheres proved to be cytocompatible and provided sustained release of the AONs for several weeks, causing increased cellular uptake in mdx dystrophic mouse cells. Here, we further investigated this delivery strategy by examining in vivo efficacy of this approach. The 2OMePS/PEI polyplexes loaded in PF microspheres were delivered by intramuscular (IM) or intra-femoral (IF) injections. We examined the carrier biodegradation profiles, AON uptake efficiency, dystrophin restoration, and muscle histopathology. Both administration routes enhanced dystrophin restoration and improved the histopathology of the mdx mice muscles. The IF administration of the microspheres improved the efficacy of the 2OMePS AONs over the IM administration. This was demonstrated by a higher exon skipping percentage and a smaller percentage of centered nucleus fibers (CNF) found in H&E-stained muscles. The restoration of dystrophin expression found for both IM and IF treatments revealed a reduced dystrophic phenotype of the treated muscles. The study concludes that injectable PF microspheres can be used as a carrier system to improve the overall therapeutic outcomes of exon skipping-based therapy for treating DMD.

Iron-chelating backbone coupled with monoamine oxidase inhibitory moiety as novel pluripotential therapeutic agents for Alzheimer's disease: a tribute to Moussa Youdim.

It is for these authors a great privilege to dedicate this review article to Moussa Youdim, who is one of the most imperative pharmacologists and pioneer investigators in the search and development of novel therapeutics for neurodegenerative diseases. 40 years ago, Moussa Youdim has started studying brain iron, catecholamine receptor and monoamine oxidase (MAO)-A and -B functions. Although Moussa Youdim succeeded in exploring the novel anti-Parkinsonian, selective MAO-B inhibitor drug, rasagiline (Azilect, Teva Pharmaceutical Co.), he did not stop searching for superior therapeutic approaches for neurodegenerative disorders. To date, Moussa Youdim and his research group are designing and synthesizing pluripotential drug candidates possessing diverse pharmacological properties that can act on multiple targets and pathological features ascribed to Parkinson's disease, Alzheimer's disease (AD) and amyotrophic lateral sclerosis. One such example is the multimodal non-toxic, brain-permeable iron-chelating compound, M30 (5-[N-methyl-N-propargylaminomethyl]-8-hydroxyquinoline), which amalgamates the propargyl moiety of rasagiline with the backbone of the potent iron chelator, VK28. This review discusses the multiple effects of several leading compounds of this series, concerning their neuroprotective/neurorestorative molecular mechanisms in vivo and in vitro, with a special focus on the pathological features ascribed to AD, including antioxidant and iron chelating activities, regulation of amyloid precursor protein and amyloid ß peptide expression processing, activation of pro-survival signaling pathways and regulation of cell cycle and neurite outgrowth.

The neuroprotective mechanism of 1-(R)-aminoindan, the major metabolite of the anti-parkinsonian drug rasagiline.

The anti-parkinsonian drug, rasagiline [N-propargyl-1-(R)-aminoindan; Azilect(R)], is a secondary cyclic benzylamine and indane derivative, which provides irreversible, potent monoamine oxidase-B (MAO-B) inhibition and possesses neuroprotective and neurorestorative activities. A prospective clinical trial has shown that rasagiline confers significant symptomatic improvement and demonstrated alterations in Parkinson's disease progression. Rasagiline is primarily metabolized by hepatic cytochrome P-450 to form its major metabolite, 1-(R)-aminoindan, a non-amphetamine, weak reversible MAO-B inhibitor compound. Recent studies indicated the potential neuroprotective effect of 1-(R)-aminoindan, suggesting that it may contribute to the overall neuroprotective and antiapoptotic effects of its parent compound, rasagiline. This review article briefly highlights the molecular mechanisms underlying the neuroprotective properties of the active metabolite of rasagiline, 1-(R)-aminoindan, supporting the valuable potential of rasagiline for disease modification.

Metallocorroles as cytoprotective agents against oxidative and nitrative stress in cellular models of neurodegeneration.

Water-soluble iron, and manganese(III) complexes of corroles and porphyrins were examined with regard to their neuroprotective/neurorescue activities by using various neuronal cytotoxic models of oxidative and nitrative stress. The present study demonstrates that the metallocorroles significantly protect human neuroblastoma SH-SY5Y and mouse motor neuron-neuroblastoma fusion NSC-34 cell lines against neurotoxicity induced by either the peroxynitrite donor 3-morpholinosydnonimine or the parkinsonism-related neurotoxin 6-hydroxydopamine. The neuronal survival effect is further reflected by the prevention of 3-morpholinosydnonimine-induced protein nitration, inhibition of caspase 3 activation, as well as attenuation of 6-hydroxydopamine-mediated decrease in growth associated protein-43 levels. The iron(III) corrole, but not manganese (III) corrole, also significantly promotes neuronal survival of hydrogen peroxide (H(2)O(2))-impaired SH-SY5Y and NSC-34 cells. A substantial superiority of the metallocorroles relative to the corresponding porphyrin complexes is revealed in all examined aspects. These results highlight the large potential of corrole complexes as novel agents for therapeutic approaches in degenerative disorders of the central and peripheral nervous systems, where oxidative and nitrative stresses are involved.

The novel multifunctional, iron-chelating drugs M30 and HLA20 protect pancreatic beta-cell lines from oxidative stress damage.

Increasing evidence suggests that oxidative stress (OS)-induced pancreatic beta-cell impairments is involved in diabetes and diabetic complications. Our group has recently synthesized two multifunctional nontoxic, lipophilic, iron-chelating drugs, 5-{N-methyl-N-propargylaminomethyl}-8-hydroxyquinoline (M30) and 5-{4-propargylpiperazin-1-ylmethyl}-8-hydroxyquinoline (HLA20), for the treatment of various OS-mediated pathogeneses. These compounds contain the N-propargylamine cytoprotective moiety of the antiparkinsonian drug rasagiline (Azilect) and the iron-complexing component 8-hydroxyquinoline. The aim of this research was to evaluate the protective effect of the multifunctional iron-chelating drugs on rat insulin-producing pancreatic beta-cells (INS-1E and RINm) against OS-induced cytotoxicity. We found that M30 and HLA20 markedly and dose-dependently inhibited H(2)O(2)-induced cytotoxicity, associated with decreased intracellular reactive oxygen species formation and increased catalase activity. In accordance, the catalase inhibitor 3-amino-1,2,4-triazol blocked the protective action of M30 against H(2)O(2)-induced damage. Both compounds significantly increased the levels of the iron-responsive protein transferrin receptor indicating their iron-chelating effect. Further mechanistic studies showed that M30 and HLA20 attenuated H(2)O(2)-induced mitochondrial membrane potential loss, decreased the release of cytochrome c into the cytoplasm, and inhibited the activation of caspase-3, suggesting that these drugs may produce cytoprotective effects via the preservation of mitochondrial function. These results indicate that the novel drugs, M30 and HLA20 display significant cytoprotective activity against OS-induced cytotoxicity in insulin producing beta-cells, which might be of therapeutic use in the treatment of diabetes mellitus.

The novel cholinesterase-monoamine oxidase inhibitor and antioxidant, ladostigil, confers neuroprotection in neuroblastoma cells and aged rats.

The current therapeutic advance in which future drugs are designed to possess varied pharmacological properties and act on multiple targets has stimulated the development of the multimodal drug, ladostigil (TV3326; (N-propargyl-(3R) aminoindan-5yl)-ethyl methyl carbamate). Ladostigil combines neuroprotective effects with monoamine oxidase (MAO)-A and MAO-B and cholinesterase (ChE) inhibitory activities in a single molecule, as a potential treatment for Alzheimer's disease (AD) and Lewy body disease. In the present study, we demonstrate that ladostigil (10(-6)-10 muM) dose-dependently increased cell viability, associated with increased activity of catalase and glutathione reductase and decrease of intracellular reactive oxygen species production in a cytotoxic model of human SH-SY5Y neuroblastoma cells exposed to hydrogen peroxide (H(2)O(2)). In addition, ladostigil significantly upregulated mRNA levels of several antioxidant enzymes (catalase, NAD(P)H quinone oxidoreductase 1 and peroxiredoxin 1) in both H(2)O(2)-treated SH-SY5Y cells, as well as in the high-density human SK-N-SH neuroblastoma cultured apoptotic models. In vivo chronic treatment with ladostigil (1 mg/kg per os per day for 30 days) markedly upregulated mRNA expression levels of various enzymes involved in metabolism and oxidation processes in aged rat hippocampus. In addition to its unique combination of ChE and MAO enzyme inhibition, these results indicate that ladostigil displays neuroprotective activity against oxidative stress-induced cell apoptosis, which might be valuable for aging and age-associated neurodegenerative diseases.

Iron dysregulation in Alzheimer's disease: multimodal brain permeable iron chelating drugs, possessing neuroprotective-neurorescue and amyloid precursor protein-processing regulatory activities as therapeutic agents.

Considering the multi-etiological character of Alzheimer's disease (AD), the current pharmacological approaches using drugs oriented towards a single molecular target possess limited ability to modify the course of the disease and thus, offer a partial benefit to the patient. In line with this concept, novel strategies include the use of a cocktail of several drugs and/or the development of a single molecule, possessing two or more active neuroprotective-neurorescue moieties that simultaneously manipulate multiple targets involved in AD pathology. A consistent observation in AD is a dysregulation of metal ions (Fe(2+), Cu(2+) and Zn(2+)) homeostasis and consequential induction of oxidative stress, associated with beta-amyloid aggregation and neurite plaque formation. In particular, iron has been demonstrated to modulate the Alzheimer's amyloid precursor holo-protein expression by a pathway similar to that of ferritin L-and H-mRNA translation through iron-responsive elements in their 5'UTRs. This review will discuss two separate scenarios concerning multiple therapy targets in AD, sharing in common the implementation of iron chelation activity: (i) novel multimodal brain-permeable iron chelating drugs, possessing neuroprotective-neurorescue and amyloid precursor protein-processing regulatory activities; (ii) natural plant polyphenols (flavonoids), such as green tea epigallocatechin gallate (EGCG) and curcumin, reported to have access to the brain and to possess multifunctional activities, such as metal chelation-radical scavenging, anti-inflammation and neuroprotection.

The neuroprotective mechanism of action of the multimodal drug ladostigil.

The recent therapeutic approach in which drug candidates are designed to possess diverse pharmacological properties and act on multiple targets has stimulated the development of the multimodal drug, ladostigil (TV3326) ((N-propargyl-(3R) aminoindan-5yl)-ethyl methyl carbamate). Ladostigil combines neuroprotective effects with monoamine oxidase -A and -B and cholinesterase inhibitory activities in a single molecule, as a potential treatment for Alzheimer's disease (AD) and Lewy Body disease. Preclinical studies show that ladostigil has antidepressant and anti-AD activities and the clinical development is planned for these dementias. In this review, we discuss the multimodal effects of ladostigil in terms of neuroprotective molecular mechanism in vivo and in vitro, which include the amyloid precursor protein processing; activation of protein kinase C and mitogen-activated protein kinase pathways; regulation of the Bcl-2 family members; inhibition of cell death markers and up-regulation of neurotrophic factors. Altogether, these scientific findings make ladostigil a potentially valuable drug for the treatment of AD.

Physiological and pathological aspects of Abeta in iron homeostasis via 5'UTR in the APP mRNA and the therapeutic use of iron-chelators.

Many studies have highlighted the pathological involvement of iron accumulation and iron-related oxidative stress (OS) in Alzheimer's disease (AD). Iron was further demonstrated to modulate expression of the Alzheimer's amyloid precursor holo-protein (APP) by a mechanism similar to that of regulation of ferritin-L and -H mRNA translation through an iron-responsive element (IRE) in their 5' untranslated regions (UTRs). Here, we discuss two aspects of the link between iron and AD, in relation to the recently discovered IRE in the 5'UTR of APP mRNA. The first is the physiological aspect: a compensatory neuroprotective response of amyloid-beta protein (Abeta) in reducing iron-induced neurotoxicity. Thus, given that Abeta possesses iron chelation sites, it is hypothesized that OS-induced intracellular iron may stimulate APP holo-protein translation (via the APP 5'UTR) and subsequently the generation of its cleavage product, Abeta, as a compensatory response that eventually reduces OS. The second is the pathological aspect: iron chelating compounds target the APP 5'UTR and possess the capacity to reduce APP translation, and subsequently Abeta levels, and thus represent molecules with high potential in the development of drugs for the treatment of AD.

TVP1022 and propargylamine protect neonatal rat ventricular myocytes against doxorubicin-induced and serum starvation-induced cardiotoxicity.

We recently reported that propargylamine derivatives such as rasagiline (Azilect) and its S-isomer TVP1022 are neuroprotective. The aim of this study was to test the hypothesis that the neuroprotective agents TVP1022 and propargylamine (the active moiety of propargylamine derivatives) are also cardioprotective. We specifically investigated the protective efficacy of TVP1022 and propargylamine in neonatal rat ventricular myocytes (NRVM) against apoptosis induced by the anthracycline chemotherapeutic agent doxorubicin and by serum starvation. We demonstrated that pretreatment of NRVM cultures with TVP1022 or propargylamine attenuated doxorubicin-induced and serum starvation-induced apoptosis, inhibited the increase in cleaved caspase 3 levels, and reversed the decline in Bcl-2/Bax ratio. These cytoprotective effects were shown to reside in the propargylamine moiety. Finally, we showed that TVP1022 neither caused proliferation of the human cancer cell lines HeLa and MDA-231 nor interfered with the anti-cancer efficacy of doxorubicin. These results suggest that TVP1022 should be considered as a novel cardioprotective agent against ischemic insults and against anthracycline cardiotoxicity and can be coadministered with doxorubicin in the treatment of human malignancies.

The neuroprotective effect of ladostigil against hydrogen peroxide-mediated cytotoxicity.

The multifunctional, anti-Alzheimer drug, ladostigil (TV3326) [(N-propargyl-(3R) aminoindan-5yl)-ethyl methyl carbamate] combines the neuroprotective effects of the anti-Parkinson drug, rasagiline, a selective monoamine oxidase (MAO)-B inhibitor, with the cholinesterase (ChE) inhibitory activity of rivastigmine in a single molecule. Ladostigil has been shown to possess potent antiapoptotic and neuroprotective activities in various oxidative insults in vitro and in vivo, such as prevention of the fall in mitochondrial membrane potential and regulation of Bcl-2 family proteins. In the present study, we demonstrate that ladostigil (1 microM) increased cell viability, associated with the increase of catalase activity and decrease of intracellular reactive oxygen species (ROS) production in human SH-SY5Y neuroblastoma cells exposed to (hydrogen peroxide) H(2)O(2). Furthermore, ladostigil significantly elevated mRNA levels of the antioxidants enzymes, catalase, NAD(P)H quinone oxidoreductase 1 (NQO1) and peroxiredoxin 1 (Prx 1) in H(2)O(2)-treated SH-SY5Y cells. Chronic treatment with ladostigil (1 mg/kg gavage per day for 30 days) markedly up-regulated mRNA expression levels of various antioxidant enzymes in aged rat hippocampus (e.g. glutathione peroxidase precursor (GSHPX-P), glutathione S-transferase (GST) and glucose-6-phosphate dehydrogenase (G6PD)). These findings indicate that in addition to its multiple neuroprotective characteristics, ladostigil also possesses antioxidant properties, which might be beneficial for the treatment of oxidative stress (OS) in aging and age-associated neurodegenerative diseases.

Induction of neurotrophic factors GDNF and BDNF associated with the mechanism of neurorescue action of rasagiline and ladostigil: new insights and implications for therapy.

Parkinson's disease (PD) and Alzheimer's disease (AD) are the most common neurodegenerative disorders, although there is no drug or therapeutic treatment to demonstrate disease-modifying effects. Previous work has proposed that neurodegeneration is linked to a lack of trophic support in those neurons and brain areas associated with PD and AD. Indeed, previous studies have found that neurotrophic factors (NTFs) support neuronal survival in various cellular and animal models of PD and AD. Thus, attention has begun to turn to the possibility of NTF neuroprotective-neurorescue therapies for these diseases, indicating that NTFs may be of significant clinical importance as exogenously supplied or endogenously induced elements that obliterate neuronal deficits and degeneration. We have recently reported that the anti-PD drug rasagiline, the anti-AD drug ladostigil, and their propargyl moiety, propargylamine, enhanced the expression levels of brain-derived neurotrophic factor and glial cell line-derived neurotrophic factor, endogenous NTFs associated with activation of phosphatidylinositol 3-kinase, protein kinase, and mitogen-activated protein kinase cell signaling/survival pathways. These studies indicate that the induction of NTFs by rasagiline and ladostigil might suppress apoptosis and induce neurorescue in neurodegenerative disorders and may support the drugs' possible disease-modifying mechanism of action.

A multifunctional, neuroprotective drug, ladostigil (TV3326), regulates holo-APP translation and processing.

The recent therapeutic approach in which drug candidates are designed to possess diverse pharmacological properties and act on multiple targets has stimulated the development of the bifunctional drug ladostigil (TV3326) [(N-propargyl-(3R) aminoindan-5yl)-ethyl methyl carbamate]. Ladostigil combines the neuroprotective effects of the antiparkinson drug rasagiline, a selective monoamine oxidase (MAO)-B inhibitor, with the cholinesterase (ChE) inhibitory activity of rivastigmine in a single molecule, as a potential treatment for Alzheimer's disease (AD) and Lewy Body disease. Here, we assessed the dual effects of lodostigil in terms of the molecular mechanism of neuroprotection and amyloid precursor protein (APP) regulation/processing by using an apoptotic model of neuroblastoma SK-N-SH cells. Ladostigil dose-dependently decreased cell death via inhibition of the cleavage and prevention of caspase-3 activation (IC50=1.05 microM) through a mechanism related to regulation of the Bcl-2 family proteins, which resulted in reduced levels of Bad and Bax and induced levels of Bcl-2 gene and protein expression. We have also followed APP regulation/processing and found that ladostigil markedly decreased apoptotic-induced levels of holo-APP protein without altering APP mRNA levels, suggesting a posttranscriptional mechanism. In addition, the drug-elevated phosphorylated protein kinase C (pPKC) levels and stimulated the release of the nonamyloidogenic alpha-secretase proteolytic pathway. Similar to ladostigil, its S-isomer, TV3279, which is a ChE inhibitor but lacks MAO inhibitory activity, exerted neuroprotective properties and regulated APP processing, indicating that these effects are independent of MAO inhibition.

Novel multifunctional anti-Alzheimer drugs with various CNS neurotransmitter targets and neuroprotective moieties.

Traditionally, drug design programs are focused on optimizing the specificity of lead compounds against a carefully selected drug target. Disappointingly, this approach to discover a "magic bullet" drug has not met with the expected success for CNS disorders. Transcriptomics and proteomic profiling of neurodegenerative diseases have indicated that they are poly-etiological in origin and that the processes leading to neuronal death are multifactorial. An emerging concept is the design of drug ligands that modulate multiple drug targets identified for a particular disease. In this review we explore some examples of multifunctional drugs which may be useful in the treatment of neurodegenerative diseases, such as Alzheimer's and Parkinson's disease.

The novel multitarget iron chelating and propargylamine drug M30 affects APP regulation and processing activities in Alzheimer's disease models.

In many of the neurodegenerative diseases, such as Alzheimer's disease (AD) and AD-related disorders, as well as in the regular ageing process, excessive generation of oxidative stress (OS) and accumulation of iron levels and deposition have been observed in specific affected-brain regions and thus, regarded as contributing factors to the pathogenesis of the diseases. In AD, iron promotes amyloid ß (Aß) neurotoxicity by producing free radical damage and OS in brain areas affected by neurodegeneration, presumably by facilitating the aggregation of Aß. In addition, it was shown that iron modulates intracellular levels of the holo amyloid precursor protein (APP) by iron-responsive elements (IRE) RNA stem loops in the 5' untranslated region (5'UTR) of the APP transcript. As a consequence of these observations, iron chelation is one of the major new therapeutic strategies for the treatment of AD. This review describes the benefits and importance of the multimodal brain permeable chimeric iron-chelating/propargylamine drug M30, concerning its neuroprotective/neurorestorative inter-related activities relevant of the pathological features ascribed to AD, with a special focus on the effect of the drug on APP regulation and processing.