Marc De Ryck

Brivaracetam: Rationale for discovery and preclinical profile of a selective SV2A ligand for epilepsy treatment

Despite availability of effective antiepileptic drugs (AEDs), many patients with epilepsy continue to experience refractory seizures and adverse events. Achievement of better seizure control and fewer side effects is key to improving quality of life. This review describes the rationale for the discovery and preclinical profile of brivaracetam (BRV), currently under regulatory review as adjunctive therapy for adults with partial‐onset seizures. The discovery of BRV was triggered by the novel mechanism of action and atypical properties of levetiracetam (LEV) in preclinical seizure and epilepsy models. LEV is associated with several mechanisms that may contribute to its antiepileptic properties and adverse effect profile. Early findings observed a moderate affinity for a unique brain‐specific LEV binding site (LBS) that correlated with anticonvulsant effects in animal models of epilepsy. This provided a promising molecular target and rationale for identifying selective, high‐affinity ligands for LBS with potential for improved antiepileptic properties. The later discovery that synaptic vesicle protein 2A (SV2A) was the molecular correlate of LBS confirmed the novelty of the target. A drug discovery program resulted in the identification of anticonvulsants, comprising two distinct families of high‐affinity SV2A ligands possessing different pharmacologic properties. Among these, BRV differed significantly from LEV by its selective, high affinity and differential interaction with SV2A as well as a higher lipophilicity, correlating with more potent and complete seizure suppression, as well as a more rapid brain penetration in preclinical models. Initial studies in animal models also revealed BRV had a greater antiepileptogenic potential than LEV. These properties of BRV highlight its promising potential as an AED that might provide broad‐spectrum efficacy, associated with a promising tolerability profile and a fast onset of action. BRV represents the first selective SV2A ligand for epilepsy treatment and may add a significant contribution to the existing armamentarium of AEDs.

Primary Amino Acid Derivatives: Compounds with Anticonvulsant and Neuropathic Pain Protection Activities

Pharmacological management remains the primary method to treat epilepsy and neuropathic pain. We have advanced a novel class of anticonvulsants termed functionalized amino acids (FAAs). In this study, we examine FAA derivatives from which the terminal acetyl moiety was removed and termed these compounds primary amino acid derivatives (PAADs). Twenty-seven PAADs were prepared; the central C(2) R-substituent was varied, including C(2) stereochemistry, and the compounds were tested in rodent models of seizures and neuropathic pain. C(2)-Hydrocarbon N-benzylamide PAADs were potent anticonvulsants and excellent anticonvulsant activity (mice, ip; rat, po) was observed for C(2) R-substituted PAADs in which the R group was ethyl, isopropyl, or tert-butyl, and the C(2) stereochemistry conformed to the d-amino acid configuration ((R)-stereoisomer). These values surpassed the activities of several clinical antiepileptic drugs. The C(2) (R)-ethyl and C(2) (R)-isopropyl PAADs also displayed excellent activities in the mouse (ip) formalin neuropathic pain model. Significantly, unlike the FAA structure-activity relationship, PAAD anticonvulsant activity increased upon substitution of a methylene unit for a heteroatom in the R-substituent that was one atom removed from the C(2) site, suggesting that these PAADs function by a different pathway than FAAs.

Synthesis, anticonvulsant activity, and neuropathic pain-attenuating activity of N-benzyl 2-amino-2-(hetero)aromatic acetamides

N-Benzyl 2-acetamido-2-substituted acetamides, where the 2-substituent is a (hetero)aromatic moiety, are potent anticonvulsants. We report the synthesis and whole animal pharmacological evaluation of 16 analogues where the terminal 2-acetyl group was removed to give the corresponding primary amino acid derivatives (PAADs). Conversion to the PAAD structure led to a substantial drop in seizure protection in animal tests, demonstrating the importance of the N-acetyl moiety for anticonvulsant activity. However, several of the PAADs displayed notable pain-attenuating activities in a mouse model.

Behavioural phenotyping reveals anxiety-like features of SV2A deficient mice.

Synaptic vesicle protein 2A (SV2A) is involved in neurotransmitter release and has been identified as the binding site for levetiracetam (Keppra), a novel antiepileptic drug. Homozygous SV2A (-/-) mice are not viable beyond a few weeks. In contrast, heterozygous SV2A (+/-) mice have a normal lifespan. We performed a behavioural phenotyping on SV2A (+/-) mice in a battery of tests: gross behavioural observation, spontaneous locomotor activity, sensori-motor coordination, acute pain sensitivity, exploration in an elevated plus-maze and an assessment of learning abilities in an inhibitory avoidance procedure. SV2A (+/-) mice were compared to age-matched, 2-month-old wild type controls. Overall, gross behaviour, spontaneous locomotor activity, sensori-motor coordination and acute pain sensitivity were comparable between wild type and SV2A (+/-) mice. When tested in a plus-maze, SV2A (+/-) mice displayed significant increased avoidance of open elevated arms whereas locomotor activity was not altered. Finally, both SV2A (+/-) and wild type mice showed comparable memory performance at the end of a multi-trial passive avoidance procedure. Interestingly, SV2A (+/-) mice exhibited increased avoidance of the lit area during the first sessions without foot shock. These results suggest an anxiety-like phenotype for SV2A (+/-) mice indicated by increased open-arm avoidance in the elevated plus-maze test as well as a shorter latency to escape from a lit area in the inhibitory avoidance procedure.

Characterization of P2X4 receptor agonists and antagonists by calcium influx and radioligand binding studies

Graphical abstract Figure. No Caption available. ABSTRACT Antagonists for ATP‐activated P2X4 ion channel receptors are currently in the focus as novel drug targets, in particular for the treatment of neuropathic and inflammatory pain. We stably expressed the human, rat and mouse P2X4 receptors in 1321N1 astrocytoma cells, which is devoid of functional nucleotide receptors, by retroviral transfection, and established monoclonal cell lines. Calcium flux assay conditions were optimized for high‐throughput screening resulting in a Z′‐factor of >0.8. The application of ready‐to‐use frozen cells did not negatively affect the results of the calcium assays, which is of great advantage for the screening of compound libraries. Species differences were observed, the rat P2X4 receptor being particularly insensitive to many ATP derivatives. Membrane preparations of the cell lines showed high levels of specific [35S]ATP&ggr;S binding with low nonspecific binding (<5% of total binding), while non‐transfected cells were devoid of specific binding sites for the radioligand. Conditions were employed which allow binding studies to be performed at room temperature. While a variety of nucleotide‐derived agonists and the antagonist TNP‐ATP displaced [35S]ATP&ggr;S from its binding site at human P2X4 receptors, the non‐nucleotidic antagonists paroxetine and 5‐BDBD did not compete with radioligand binding and were therefore characterized as allosteric antagonists. Homology modeling was applied to find an explanation for the observed species differences.

UCB Pharma research day-25 October 2007 'Glia-neuron interactions and purinergic receptors in neurological disorders'.

UCB Pharma asked Geoffrey Burnstock to propose a programme of 8 international leaders in the glia-neuron interaction and purinergic signalling field to participate in a meeting aimed at introducing the topic to about 30 members of the Company to consider the therapeutic potential of purinergic signalling compounds for the treatment of neurological disorders, including: epilepsy, neurodegenerative diseases, movement disorders, neuroprotection, mood disorders and neuropathic pain.

Discovery of selective alpha(2C) adrenergic receptor agonists.

Neuropathic pain has recently been defined by the International Association for the Study of Pain (IASP) as “pain caused by a lesion or disease of the somatosensory system”. Neuropathic pain is chronic and may involve the central nervous system (CNS) (e. g. , stroke-related pain), as well as the peripheral nervous system (e. g. , trauma, resection, infection or progressive degeneration of a peripheral nerve). Examples of neuropathic pain include diabetic painful neuropathy, post-herpetic neuralgia, trigeminal neuralgia, cancerand HIV-related neuropathic pain, and iatrogenic neuropathic pain, associated with postsurgical neuropathy and neuropathies induced by cancer chemotherapy or antiretroviral drugs. Currently marketed drugs treating neuropathic pain are not optimal since they alleviate pain in only about 50 % of patients, making the treatment of neuropathic pain an unmet medical need. 4] Anti-epileptic drugs, such as gabapentin (Neurontin) and pregabalin (Lyrica), are the most commonly prescribed treatment. Delineating the specific role of different neurotransmitter–receptor systems in conducting pain messages has been the subject of intense research efforts. In particular, monoaminergic neurotransmitters, including serotonin, dopamine and noradrenaline (NA), are considered as mechanistic targets for pain. For instance, Tanabe and co-workers have provided evidence that gabapentin is characterized by a supraspinal site of action, in which central NA is involved. Their results indicate that gabapentin acts on supraspinal structures to activate the descending noradrenergic system. This terminates in the lumbar spinal cord, where NA interacts with a2 adrenergic receptors (ARs) to reduce the transmission of nociceptive information. The role of the noradrenergic system in pain mechanisms was confirmed when the drug clonidine (1), an a2-AR agonist and initially designed to reduce blood pressure in patients with essential hypertension, demonstrated marked anesthetic and analgesic properties. Clonidine became the first drug to be approved by the US Food and Drug Administration (FDA) for the treatment of cancer-related neuropathic pain as a substitute for opioids in patients suffering from excessive tolerance-related side effects. However, clonidine produces clinically relevant, dose-limiting adverse effects, such as arterial hypotension and sedation, that negatively impact its use, which, in cancer neuropathy, is limited to spinal infusion. The neurotransmitter NA acts through several ARs, including a2-ARs belonging to the seven-transmembrane G protein-coupled receptors (GPCRs). The a2-ARs are subdivided into a2A/D, a2B and a2C subtypes, [9–11] where a2A/D-AR is a global nomenclature for the human a2Aand murine a2D-AR orthologues. [12]