Advances toward precision therapeutics for developmental and epileptic encephalopathies

Abstract

Developmental and epileptic encephalopathies are childhood syndromes of severe epilepsy associated with cognitive and behavioral disorders. Of note, epileptic seizures represent only a part, although substantial, of the clinical spectrum. Whether the epileptiform activity per se accounts for developmental and intellectual disabilities is still unclear. In a few cases, seizures can be alleviated by antiseizure medication (ASM). However, the major comorbid features associated remain unsolved, including psychiatric disorders such as autism-like and attention deficit hyperactivity disorder-like behavior. Not surprisingly, the number of genes known to be involved is continuously growing, and genetically engineered rodent models are valuable tools for investigating the impact of gene mutations on local and distributed brain circuits. Despite the inconsistencies and problems arising in the generation and validation of the different preclinical models, those are unique and precious tools to identify new molecular targets, and essential to provide prospects for effective therapeutics.

Keywords: adeno-associated virus; epilepsy; rare diseases; rodent models; therapeutics.

Figure 1

Etiology and circuit-based malfunctions in epilepsy. (A) Different genes, environmental/epigenetic factors and brain injuries may be the cause of circuit malfunctions that give rise to epilepsy, a network disease involving different interacting brain regions; (B) the prevalence of epileptic diseases is classified into common and rare epilepsies (~95 and ~5%, respectively). Developmental and epileptic encephalopathies (DEEs) are rare diseases mainly occurring in pediatric age. (C) In epileptic disorders the excitatory drive increases, generating hyperexcitability and E/I dysbalance; (D) epilepsy is initiated in different brain regions and activates synaptic brain circuits for seizure maintenance. It can further spread to other brain regions, generating comorbidities and increasing seizure severity. (E) Genetic rodent models for DEE mainly consist of knockout, multigenic deletions, and knock-in. These latter may be based on precise human point mutations, satisfying the criteria for ‘construct validity completely’; (F) Different pros and cons characterize these animal models. The most significant caveats concern the frequent inconsistencies with the human phenotype, particularly the possible absence of the epileptic phenotype and the incomplete penetrance of the mutation. The few models showing spontaneous epileptic seizures are often not viable or affected by a high mortality rate, and the seizures’ unpredictability makes them difficult to study. Despite this, genetic animal models are unique and valuable tools for assessing new candidate ASM and other therapeutic strategies and are necessary for subsequent clinical studies, thus benefits prevail.

Figure 2

Inducible control of excitation-inhibition balance and gene activity. (A) Animal models are useful for testing specific hypotheses-driven experiments, also regarding gender differences in various pathologies and in response to therapy. Thus far, they represent a powerful approach to move preclinical studies into clinical trials; (B) Recombinant adeno-associated virus (rAAVs) can be equipped with α-calmodulin kinase-2 promoter (αCaMK2) to express reverse tetracycline transactivator (rtTA; rAAV1), which upon (C) binding to the Ptetbi promoter (rAAV3) in the presence of doxycycline (Dox) activate the expression of tetanus toxin light chain (TeTxLC) to block evoked synaptic transmission in excitatory neurons. In the absence of Dox, the expression is switched-off and synaptic transmission can resume; (D) With another system based on c-fos promoter with upstream tetracycline operators (t), hyperactive cells are specifically targeted in the presence of Dox to express rtTA, which establishes an autoregulated loop for rtTA-dependent rtTA expression, thereby E/I can be controlled in the tagged cells using TeTxLC or any gene of interest (GOI) as in (C). Indeed, these technologies shown here (B/C) or (D/C) can also be used to express any gene of interest to override the mutations responsible for epilepsy. (E) Moreover, the level of expression and synaptic silencing can be controlled at different concentrations of Dox. Controlled levels of gene expression can allow for proper titration of E/I balance.