European Journal of Paediatric Neurology 47 (2023) 25–34

Contents lists available at ScienceDirect

European Journal of Paediatric Neurology

journal homepage: www.journals.elsevier.com/european-journal-of-paediatric-neurology

Updated clinical recommendations for the management of tuberous

sclerosis complex associated epilepsy
Nicola Specchio a, *, Rima Nabbout b, Eleonora Aronica c, d, Stephane Auvin e, f, g,
Arianna Benvenuto h, Luca de Palma a, Martha Feucht i, 1, Floor Jansen j, 2, Katarzyna Kotulska k, 3,
Harvey Sarnat l, Lieven Lagae m, Sergiusz Jozwiak n, 3, Paolo Curatolo o
a
Clinical and Experimental Neurology, Bambino Gesu’ Children’s Hospital IRCCS, Full Member of European Reference Network on Rare and Complex Epilepsies
EpiCARE, Rome, Italy
b
Department of Pediatric Neurology, Necker Enfants Malades Hospital, Université Paris Cité, Member of the European Reference Network on Rare and Complex
Epilepsies EpiCARE, INSERM U1163, Institut Imagine, Paris, France
c
Amsterdam UMC, University of Amsterdam, Amsterdam Neuroscience, Department of (Neuro)Pathology, Amsterdam, Netherlands
d
Stichting Epilepsie Instellingen Nederland (SEIN), Heemstede, the Netherlands
e
APHP, Service de Neurologie Pédiatrique, Centre Epilepsies Rares, Member of the European Reference Network on Rare and Complex Epilepsies EpiCARE, Hôpital
Robert Debré, Paris, France
f
Université Paris-Cité, INSERM NeuroDiderot, Paris, France
g
Institut Universitaire de France (IUF), Paris, France
h
Libera Università Maria Santissima Assunta (LUMSA), Rome, Italy
i
Epilepsy Center, Department of Pediatrics, Medical University Vienna, Austria
j
Department of Pediatric Neurology, Brain Center UMC Utrecht, the Netherlands
k
Department of Neurology and Epileptology, The Children’s Memorial Health Institute, Warsaw, Poland
l
Department of Paediatrics (Neurology), Pathology and Laboratory Medicine (Neuropathology) and Clinical Neurosciences, University of Calgary Cumming School of
Medicine and Alberta Children’s Hospital Research Institute (Owerko Centre), Calgary, AB, Canada
m
Department of Paediatric Neurology, University of Leuven, Leuven, Belgium
n
Research Department, The Children’s Memorial Health Institute, ERN EPICARE, Warsaw, Poland
o
Child Neurology and Psychiatry Unit, Systems Medicine Department, Tor Vergata University, Rome, Italy

A R T I C L E I N F O A B S T R A C T

Keywords: Children with tuberous sclerosis complex (TSC), may experience a variety of seizure types in the first year of life,
Everolimus most often focal seizure sand epileptic spasms. Drug resistance is seen early in many patients, and the man­
Epileptogenesis agement of TSC associated epilepsy remain a major challenge for clinicians. In 2018 clinical recommendations
Developmental and epileptic encephalopathies
for the management of TSC associated epilepsy were published by a panel of European experts. In the last five
Neurodevelopmental disorders
Pre-symptomatic treatment
years considerable progress has been made in understanding the neurobiology of epileptogenesis and three
Drug resistant epilepsy interventional randomized controlled trials have changed the therapeutic approach for the management of TSC
associated epilepsy. Pre-symptomatic treatment with vigabatrin may delay seizure onset, may reduce seizure
severity and reduce the risk of epileptic encephalopathy. The efficacy of mTOR inhibition with adjunctive
everolimus was documented in patients with TSC associated refractory seizures and cannabidiol could be another
therapeutic option. Epilepsy surgery has significantly improved seizure outcome in selected patients and should
be considered early in all patients with drug resistant epilepsy.
There is a need to identify patients who may have a higher risk of developing epilepsy and autism spectrum
disorder (ASD).
In the recent years significant progress has been made owing to the early identification of risk factors for the
development of drug-resistant epilepsy.
Better understanding of the mechanism underlying epileptogenesis may improve the management for TSC-
related epilepsy. Developmental neurobiology and neuropathology give opportunities for the implementation

* Corresponding author. Clinical and Experimental Neurology, Bambino Gesu’ Children’s Hospital, IRCCS, Piazza S. Onofrio 4, 00165, Rome, Italy.
E-mail address: nicola.specchio@opbg.net (N. Specchio).
1
Full Member of European Reference Network EpiCARE.
2
Full member of the European Reference Network on Rare and Complex Epilepsies EpiCARE.
3
Member of ERN EPICARE.

https://doi.org/10.1016/j.ejpn.2023.08.005
Received 8 March 2023; Received in revised form 27 August 2023; Accepted 28 August 2023
Available online 30 August 2023
1090-3798/© 2023 The Authors. Published by Elsevier Ltd on behalf of European Paediatric Neurology Society. This is an open access article under the CC BY-NC-ND
license (http://creativecommons.org/licenses/by-nc-nd/4.0/).
N. Specchio et al.
of concepts related to clinical findings, and an early genetic diagnosis and use of EEG and MRI biomarkers may
improve the development of pre-symptomatic and disease-modifying strategies.
1. Introduction
TSC is a multisystem genetic disorder characterized by age-related
development of benign tumors across several organs including the
brain, skin, kidneys, heart, and eyes. TSC is caused by inactivating
mutations in either TSC-1 or TSC-2 genes [1–3]. The gene mutation
results in an overactivation of the PI3K-Akt-mechanistic (formerly
mammalian) target of the rapamycin (mTOR) signaling pathway, which
plays a crucial role from brain development processes to aging [4].
Overall, more than 2000 pathogenic TSC1/2 variants have been
described in the Leiden Open Variation Database (www.lovd.nl/TSC1
and www.lovd.nl/TSC2), making the genotype – phenotype correla­
tion challenging. Identification of a pathogenic variant or deletion in
TSC1/2 is sufficient for the diagnosis of TSC, regardless of clinical
findings (Northrup 2021) [5]. However, about 5–8% of TSC individuals
meeting definite clinical diagnostic criteria have no pathogenic TSC
gene variant or deletion identified by direct sequencing of both genes or
by whole exome sequencing analysis [5–8].
Epilepsy is the most common (80–90% of patients) [9] of a large
spectrum of neurological and psychiatric manifestation in patients with
TSC. In addition neurodevelopmental disorders with intellectual
disability and autism spectrum disorder (ASD), and behavioral abnor­
malities are highly prevalent [10,11].
Patients with TSC may experience a variety of types of seizures:
epileptic spasms, focal, tonic, atonic, and tonic-clonic seizures in the
context of an evolution toward Lennox-Gastaut syndrome. The majority
of patients experience seizures in the first year of life [12,13]. About 5%
of children develop epilepsy as newborns, often due to malformations of
cortical development meeting the criteria of focal cortical dysplasia
(FCD) [14]. The recognition of subtle focal seizures in infants by care­
givers may be difficult [15].
In 2017, a panel of European specialists met in Rome to define
clinical recommendations for the management of epilepsy associated
with TSC [16]. In the last five years, considerable progress has been
made in understanding various aspects of this disorder, including the
neurobiology of (mTOR dependent) epileptogenesis.
First of all the International ‘TuberOus SClerosis registry to increase
disease Awareness’ (TOSCA) [17] has provided baseline data on more
than 2000 patients and new insights into the clinical characteristics and
the natural history of TSC. Moreover, three international randomized
controlled trials (RCTs) have changed clinical practices in the treatment
of epilepsy associated with TSC. The EPISTOP (NCT02098759) trial
showed that pre-symptomatic treatment with vigabatrin may modify the
natural history of seizures with a delay in seizure onset and a reduction
in seizure severity and the risk of epileptic encephalopathy (EE) [18].
The EXIST3 trial demonstrated the efficacy of mTOR inhibition with
adjunctive Everolimus in reducing seizure frequency in patients with
TSC-associated refractory seizures [19]. Finally, the GWPCARE6 trial
documented efficacy and safety of Cannabidiol (CBD) in reducing
TSC-associated seizures compared with placebo [20]. CBD is now
approved by Food and Drug Administration (FDA) and European Med­
icine Agency (EMA) for the treatment of seizures in patients with TSC.
Experts are now moving away from the traditional therapeutic
approach with antiseizure medication (ASM) applied after seizure onset,
to a more proactive approach focused on pre-symptomatic treatment
with vigabatrin or molecular targeted treatment with mTOR inhibition
[21]. Advances in surgery improves access to this treatment for patients
with multiple tubers [22].
To review new insights from the last years, a new panel of experts
met in Rome in November 2022 to address specific questions. Each panel
expert was assigned a topic to review beforehand and present at the
26
European Journal of Paediatric Neurology 47 (2023) 25–34
meeting. The panel addressed the mechanisms of epileptogenesis, the
predictive value of clinical EEG, brain MRI and genetic biomarkers, and
evaluated the potential benefits of pre-symptomatic treatment with
vigabatrin and targeted treatment with Everolimus, and other new
treatment possibilities, including CBD and indications for presurgical
evaluation.
This paper outlines the issues discussed at the consensus meeting
workshop and sets out updated clinical recommendations for current
treatment of TSC-associated epilepsy, considering new concepts and
changes since the last publication of the 2018 clinical recommendations.
2. Progress in understanding mTOR dependent epileptogenesis
and co-occurring neurodevelopmental disorders
TSC is genetically complex because it involves both germline genes
and somatic second-hit events. Second-hit loss of either TSC1 or TSC2,
inducing mTOR hyperactivation, has been identified in TSC neoplastic
lesions, but is less frequently detected in cortical tubers, suggesting a
role for additional molecular mechanisms [23,24].
Extensive research over the past decade has provided strong evi­
dence of the critical role of the mTOR pathway during brain develop­
ment, controlling many basic cellular functions, such as energy
metabolism, protein synthesis and autophagy, to regulate cell growth,
proliferation, migration, dendritic formation and axon elongation [25,
26]. The mTOR signaling pathway has a crucial function in the process
of neuronal and astrocytes differentiation, as well as in oligodendrocyte
lineage development. Balanced spatiotemporal mTOR signaling is
crucial for the proper development of the human cortex [25,27,28].
Accordingly dysregulation of the mTOR signaling represents a common
pathogenic mechanism of a spectrum of malformations of cortical
development, sharing histopathological and clinical features, including
epilepsy and co-occurring neurodevelopmental disorders [25,29].
Timing of mTOR activation plays a crucial role, and subsequently de­
termines the neuropathological and clinical phenotype [29–31]. Inter­
estingly, in experimental models, the level of mTOR-hyperactivity
correlates with the severity of epilepsy and associated neuropathology
[32].
Cortical tubers, subependymal nodules and subependymal giant cell
astrocytoma are the main pathologies in the brain of TSC patients.
Cortical tubers represent the neuropathological basis of epilepsy and
consist of areas of cortical dyslamination containing different cell types,
including dysmorphic neurons, giant cells and reactive astrocytes [10].
Cortical tuber formation is a complex and dynamic process starting in
fetal brain but continuing as the brain matures [10,33]. Visual detection
of cortical tubers on neuroimaging is limited in pre-natal and early
post-natal period, and it improves with the myelination process during
the first three years of life [34]. Prenatal brain MRI revealed cortical
tubers in 20 out of 30 (66.7%) patients in the study of Slowinska et al.
[35]. Predefined advanced imaging protocols with 3D T1 and T2 se­
quences are required to improve lesion detection in the new-born with
TSC. Some MRI sequences as arterial spin labelling (ASL) have been
reported of help in early childhood to identify tubers in relation to EEG
slow waves [36], but this application needs further validation.
Insights from a variety of in vivo and in vitro genetic models and from
human pathology have shown that epileptogenesis in TSC is a chronic
and dynamic process leading to early onset and difficult to treat seizures,
co-occurring neurodevelopmental disorders and mTOR-related neuro­
degeneration (i.e. increases risk for tauopathy) [10,37]. This process
may start prenatally and can continue long after the epilepsy diagnosis.
Knowledge about the complex biological mechanisms contributing to
network dysfunction during brain development, even beyond the early
N. Specchio et al. European Journal of Paediatric Neurology 47 (2023) 25–34

period of epileptogenesis, in individuals with TSC, is rapidly increasing Table 1

[10,38]. Experimental and human studies provide evidence of early and Predictors of drug resistance.
sustained oxidative stress and inflammation, accompanied by blood Predictors of drug resistance
brain barrier (BBB) dysfunction and extracellular matrix remodelling
Clinical
[10,38–40]. seizures
Dysregulation of mTOR signaling itself may result in developmental - ES + Nabbout R et al., 2019 [17], Jeong A
alterations of the balance between excitation and inhibition. Converging et al., 2017 [57]
evidence suggest that the pathological network underlying early - Focal seizures + ES +++ Nabbout R et al., 2019 [17], Jeong A
et al., 2017 [57]
appearance of seizures is characterized by persistence of immature - Younger age at onset of +++ Capal JK et al., 2017 [49]
patterns of neurotransmitter receptor expression and function (i.e. clinical seizures
GABAA-receptor [41]) and/or disrupted GABAergic interneuron EEG
network development and organization [10,38,42]. Moreover, the Interictal abnormalities
- Focal De Ridder J et al., 2020 [65]
direct influence of mTOR pathway dysregulation on oligodendrocyte +++
- Multi-focal +++ De Ridder J et al., 2020 [65], De Ridder
differentiation and myelin formation could further contribute to J et al., 2021 [62]
abnormal cell-signaling and network dysfunction [10,38,43]. Other - Younger age at onset of +++ De Ridder J et al., 2021 [62], Wu JY
pathways, that might be related to the mTOR pathway, have also been recording of IEDs et al., 2019 [59]
suggested as interesting targets in therapy of TSC-related seizures during Genetic
- TSC 2 gene variant Ogórek B et al., 2020 [60]
brain development such as the inflammatory pathway (IL-1-CXCL10)
+++

[44] and the GluN2C subunit of the NDMA receptor [45,46]. Combi­
nation of multiple targeted interventions might be needed to further
increase the efficacy of mTOR inhibitor treatment in TSC-related sei­
zures in humans.
There is a complex relationship between epilepsy and autism, and
TSC is among the major causes of syndromic ASD [11,47]. TSC2 muta­
tions, tuber volumes and location, infantile spasms and early onset sei­
zures have been reported as the most important risk factors for ASD
[48]. A developmental disruption of GABAergic cortical interneurons
linked to abnormal generation, migration routes as well as their mo­
lecular and functional diversifications can explain the concomitant
occurrence of epilepsy and autism [10,38,42]. Although there is a strong
association between the presence of epilepsy and TSC-associated ASD,
ASD should not be considered solely as a co-morbidity of early onset
epilepsy and the infantile epileptic spasms syndrome (IESS).
Ever since the last century, clinicians have always observed that
seizures appear in the first months of life, while ASD risk is recognized
much later and is typically not diagnosed before 2–3 years of age.
Therefore, their understanding was that early epilepsy onset may lead to
autism even if there is morphological substrate of autism (brain lesions)
and some autism features are seen in patients without epilepsy [11,47].
Recent evidence showed that early risk of autism can be detected
already in the first year of life [49,50]. Complex cellular and molecular
events play a significant role in determining a high risk for both epilepsy
and neurodevelopmental deficits in infants and children with TSC
[51–53]. TSC can be considered a model of genetic-determined devel­
opmental and epileptic encephalopathy, a group of severe epilepsies that
are characterized by - often drug resistant - seizures and encephalopathy
related to the dual impact of the seizures and of the etiology [54,55].
3. Predictors of drug resistance
TSC-associated epilepsy management is still a challenge because of
high percentage of patients developing drug resistant epilepsy (DRE).
Data from the TOSCA registry revealed that patients presenting with
focal seizures (whether or not combined with IESS) were more re­
fractory than patients presenting with epileptic spasms only, but the
prediction of DRE is still challenging [17].
In a prospective study, infants with an early seizure onset continued
to perform worse at 24 months of age, in comparison with children with
a later seizure onset [56]. Data from the TSC Natural History Database,
including 1965 subjects with TSC, showed that early focal-onset seizures
and treatment-resistant infantile spasms were clinical predictors for DRE
[57]. Infantile spasms were also among strong indicators of DRE as re­
ported in a systematic TSC review [58]. In a longitudinal observational
study of 40 infants interictal epileptiform discharges (IEDs) correctly
identified future epilepsy in 77% of seizure naïve infants [59]. However,
due to the high prevalence of epilepsy in TSC patients (80–90%) we
MRI
- Higher tuber load +++ Hulshof HM et al., 2022 [66]
- Tuber location + Cohen AL et al., 2021 [67]
ES: epileptic spasms, IEDs: interictal epileptiform discharges, R: right.
cannot confirm the predictive value of IEDs. The EPISTOP study has
recently identified useful pre-symptomatic genetic, EEG and MRI bio­
markers that can be used in clinical practice to identify infants at risk for
developing DRE at the age of two years [18,60–63].
TSC2 pathogenic variants were associated with a higher tuber load
on MRI and were predictive of more severe epilepsies with higher risk of
drug resistance [64]. Interictal focal or multifocal discharges on serial
sleep-wake EEG in pre-symptomatic infants with TSC are predictive of
an ongoing severe epilepsy [59]. Ninety-five per cent of infants with TSC
enrolled in the EPISTOP trial developed IED at a medium age of 77 days,
and earlier recording of IED was associated with an increased risk of
DRE [62,65]. Although fetal MRI brain lesions did not predict the risk of
clinical seizures or DRE at the age of two years [34], higher fetal lesion
load, was significantly associated with lower developmental outcome.
Furthermore, higher tuber volume as a proportion of total brain volume
on early postnatal MRI was associated with clinical seizures and DRE at
the age of two years [66].
In a prospective observational cohort of 123 infants, tuber location
was heterogeneous, however tubers functionally connected to the globi
pallidi and cerebellar vermis were stronger predictor of spasms [67].
Increased EEG connectivity was strongly associated with epileptic
spasms in another cohort [68].
In summary early recognition of pathogenic variants and the use of
reliable EEG and MRI biomarkers could help to identify pathogenic
mechanisms for epileptogenesis and provide opportunities for pre-
symptomatic and disease modifying approaches. Clinical, EEG, MRI
and genetic predictors of DRE are summarized in Table 1. Development
of a prediction model that incorporates a variety of risk factors, as well
as machine learning models with multimodality features could aid in
guidance and treatment of TSC patients with high risk of DRE or neu­
rodevelopmental disorders [69,70].
4. Pre-symptomatic treatment
Pre-symptomatic treatment with VGB was already suggested by the
previous expert panel in 2018, but its efficacy is now strongly confirmed
by the EPISTOP trial [18]. The EPISTOP trial was carried out from 2014
to 2018 in 9 European centers and one Australian site. The aim of the
trial was to evaluate the safety and efficacy of conventional versus
“preventive” treatment of epilepsy with vigabatrin at the dosage of at
least 100 mg/kg/day. Eligible patients were newborns and infants under

27
N. Specchio et al. European Journal of Paediatric Neurology 47 (2023) 25–34

Fig. 1. This is a 10-month-old boy with a TSC2 genetic variant. The diagnosis was suspected prenatally due to the presence of cardiac rhabdomyomas and confirmed
at 1 months of age, when brain MRI showed several tubers. At the age of 3 months, EEG monitoring was started (A) showing during sleep a normal background
activity with rare slow waves over the right temporal region. (B) 2 months later (age 5 months) spike and spikes and wave complexes appeared during sleep over the
left temporal region. Pre-symptomatic treatment with vigabatrin at the dose of 75 mg/kg day was started. (C) Another 2 and 4 months later (age 7 and 9 months
respectively) sleep EEG was and remained (D) normal. He did not experience clinical seizures, and the development is normal.

Table 2
Ongoing trials on prevention of epilepsy in TSC patients.
Ongoing trials

Trial Phase Intervention Study Population Primary Outcome(s) Completion

PREVeNT IIb Vigabatrin vs Placebo Infants with TSC aged 1 day to N= Cognitive assessment scores at 24 months May 2023
6 months 84
TSC- II Sirolimus vs Placebo N= % Reporting severe or serious AEs; time to seizure onset June 2026
STEPS 64 at 12 months of age
ViRap II/III Sirolimus vs Vigabatrin (masked Infants with TSC aged 4–16 N= Occurrence of clinical seizures; volume of TSC-associated Mar 2026
with placebo) weeks 60 tumors at 730 days

the age of 4 months, before the onset of seizures. All participants were
monitored with serial EEG and vigabatrin was introduced either when
IED were detected or after the onset of seizures. Preventive treatment
with vigabatrin in 25 children resulted in delay of clinical seizures,
reduced risk of developing drug resistant epilepsy, and absence of in­
fantile spasms, when compared with 25 patients taking treatment after
seizure onset [18] (Fig. 1).
In the EPISTOP study treatment with vigabatrin in TSC infants was
safe and modified the natural history of seizures, reducing the risk and
severity of epilepsy. Since epilepsy was not prevented in the majority of
infants, the term pre-symptomatic treatment is to be preferred and is
now considered an established approach in several European country in
infants with TSC [71]. Supporting this more proactive use of Vigabatrin
for TSC-epilepsy, new data published from the US Vigabatrin registry
that followed >9000 patients on Vigabatrin for up to 16 years did not
find that any patient experienced clinical vision loss, few (2%) patients
experienced new vision findings over time and none were determined to
be caused by Vigabatrin. Thus, the risk to vision appears lower than
previously thought [72]. Furthermore, early and frequent EEG moni­
toring and early (before any type of seizures, after electrographic
seizure, or immediately after first clinical seizure) treatment resulted in
better neurodevelopmental outcome than in any other historical cohort
of TSC patients (median IQ within the normal range in the whole
cohort), however, longer follow up is warranted before firm conclusions
can be drawn with respect to neurodevelopmental outcomes [18].
Autistic features were detected irrespectively of treatment strategy,
suggesting either independent mechanisms or the impact of earlier
epileptogenesis-related events since intrauterine life [50,53].
Ongoing RCTs, with the aim to prevent epilepsy, are summarized in
Table 2.
5. Targeted treatment with mTOR inhibitors
mTOR inhibition can reduce structural abnormalities in the brain:
mTOR inhibition reduces neuronal morphology abnormalities, cell size
and inflammatory mediators in the cortex, and improves neuronal
myelination and synaptic plasticity.
Initially developed as anti-fungal agents and later discovered to have
potent immunosuppressive and antiproliferative properties, rapamycin
(Sirolimus) has been found to be a mechanistic target of mTOR

28
N. Specchio et al.
overactivation [73]. Rapamycin is a macrolide compound, isolated for
the first time in 1972, from samples of Streptomyces hygroscopicus
found on Easter Island. A mechanistic target of rapamycin kinase
(mTOR) inhibitor acts on T and B cells by reducing their sensitivity to
interleukin-2 (IL-2). The compound was approved by the FDA in 1999 as
immunosuppressant indicated for the prophylaxis of organ rejection in
patients aged ≥13 years receiving renal transplants and patients with
lymphangioleiomyomatosis. Rapamycin/Sirolimus changes microglial
polarity in TCS mice [74]. It has also been shown to decrease seizures
and neuropsychiatric symptoms, except for ASD, (and this effect seems
to be related to the regulation of microglia polarity) in TSC mouse
models [75–77]. Recent uncontrolled studies demonstrated a favorable
safety and efficacy profile of sirolimus for treating manifestations of TSC
(including epilepsy); the study reported on efficacy and safety of rapa­
mycin in DRE in 32 children aged 11 months to 14 years, at least 50%
reduction of seizures was seen in 56% of patients [78]. However,
long-term efficacy/safety data from prospective controlled studies in
TSC are only available for everolimus.
Everolimus (the 40-O-(2-hydroxyethyl) derivative of Sirolimus), is a
second-generation rapamycin derivative and works similarly to siroli­
mus as an inhibitor of mTOR pathway.
Although having a similar structure everolimus significant differs
regarding its pharmacokinetic, pharmacodynamic, and toxic properties
resulting in distinct clinical profile. Everolimus is more readily absor­
bed, exhibits a greater oral bioavailability and is a selective inhibitor of
mTORC1 with little impact on mTORC2. Therefore, it could achieve
many of the benefits of sirolimus on cell growth and proliferation
without glucose intolerance [79].
There are still ongoing efforts to limit the non-neurologic effects of
mTOR inhibitors. Indeed, experimental study showed that brain-specific
analogs of rapamycin (termed rapalogs) have recently been developed by
inhibiting brain mTOR while sparing mTOR activity in other tissues
using the brain-permeable mTOR inhibitor RapaLink-1 and the brain-
impermeable FKBP12 ligand RapaBlock [80].
Everolimus, has been considered as an add-on treatment option for
individuals with TSC associated drug resistant epilepsy. Treatment ef­
ficacy and safety were reported in the EXIST-3 trial where adjunctive
everolimus achieved a clinically meaningful reduction of seizure fre­
quency with a favorable risk-benefit ratio [19,81]. Furthermore,
response to everolimus tends to improve with ongoing treatment and the
post authorization safety study (PASS) confirmed the long-term safety of
everolimus in the real-world clinical practice, and a better response for
the <6 years subgroup was documented [82,83]. The target drug con­
centration of 5–9 ng/ml was recommended [81].
Everolimus has been also studied as the treatment of TSC-associated
neuropsychiatric disorders in a RCT study in patients aged 6–21 years
comparing oral everolimus (n = 32) vs placebo (n = 15) taken once daily
for 6 months. Comprehensive neurocognitive and behavioral evaluation
battery was administered at baseline, 3 months, and 6 months. At the
end of the study everolimus did not significantly improve neuro­
development or behavior in children with TSC, possibly related to small
sample size, study design and outcomes chosen [84]. However, the panel
believe that definitive conclusion could not be provided for the efficacy
of everolimus on cognition, considered that the drug was used in in­
dividuals with older than 2 years of age with already significant neu­
rodevelopmental deficits. Recent evidence showed that everolimus is
safe also in young children under the age of 2 years [85].
Since functional connectivity is important for neurodevelopment,
the longitudinal effects of everolimus were studied on callosal white
matter diffusion tensor imaging (DTI) in patients with TSC. mTOR
overactivity on white matter microstructural integrity in TSC were
modified through pharmacologic inhibition of mTOR [86]. However,
how this effected neurodevelopment was not studied. Everolimus was
used in a preclinical model of TSC induced by Tsc2+/− , and it led to a
persistent improvement of the social deficit while deficits related to
developmental status epilepticus did not respond to treatment [87].
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European Journal of Paediatric Neurology 47 (2023) 25–34
What is still not known is the optimal timing for mTOR inhibition,
and its applicability in fetal life. Everolimus was used in a pregnant
woman to treat her fetus with TSC and regression and subsequent sta­
bilization of the cardiac and brain lesions were seen; postnatally, the
child did not develop seizures or autism and presented good neuro­
developmental outcome [88,89].
Since mTOR inhibitors may have effects on various systemic mani­
festation of TSC, the panel suggest that everolimus should be considered
early when growing subependymal giant cell astrocytoma (SEGA) or
angiomyolipoma (AML) are seen in combination with DRE [90]. Sup­
porting evidence for the efficacy of combination treatment with siroli­
mus and everolimus in treating the same/different TSC manifestations
should be available, as there are currently no head-to-head published
studies. It is important to demonstrate if dual targeting of TORC1/­
TORC2 provides superior efficacy, above sirolimus or everolimus as
monotherapy, without significant increases in toxicity. In addition, and
in contrast to oncology, sirolimus in TSC patients is used in monotherapy
in young patients and therefore might have fewer side effects.
6. Surgery: relevance and limitations
In 2017’s workshop, the panel was already suggesting that surgery
can be a good therapeutic option for TSC in a well selected population of
patients with recommendation of early presurgical evaluation to
attempt to improve neurodevelopmental outcome as well. New evidence
emphasizes and strengthens this recommendation. Overall post-
operative seizure freedom is achieved in 50–60% of TSC patients, and
>50% seizure reduction is seen in another 15%, even in patients with
multiple tubers. In selected series it has been observed that duration of
follow up matters as there is a substantial decrease of seizure-free pa­
tients with a longer follow-up [91].
Epilepsy surgery in TSC is still underused [92]. Indication have long
been limited to clear unifocal epilepsy, but the great majority of patients
have multifocal EEG abnormalities and numerous tubers. Thus, indica­
tion may need to be widened taking the high burden of predominant
seizures into account, especially when semiology is consistent over time.
A recent nationwide multicenter retrospective study in 364 patients
who underwent surgery at the median age of 8.5 years, reports sustained
seizure freedom in 51% at 10 years, particularly after complete removal
of a large lesion on MRI or after tuberectomy plus cortectomy, meaning
the epileptogenic zone beyond tuber borders [93]. A systematic review
of 46 studies regarding the outcome of 1157 patients showed an excel­
lent seizure reduction in 59% of the individuals who underwent epilepsy
surgery [91]. Very early onset epilepsy, the absence of a tuber which is
expected to predominate in generating epileptiform abnormalities and
seizures, and continuing spasms are probably related to worse outcome.
Developmental progress, but very limited improvement in the stan­
dard scales, was reported in a subset of individuals who showed a
worthwhile reduction of seizures frequency [94]. Altogether, more in­
formation on causes of surgical failures and predictors of efficacy is
needed [22,95,96] but importantly, patient satisfaction even in patients
who did not achieve seizure freedom, was high [97]. Laser ablation was
performed in three patients with promising results, but more data is
needed to recommend its use in TSC patients [98].
The current relevance and limitation of surgery for TSC-associated
drug resistant epilepsy was widely discussed by the panel. Presurgical
evaluation should be considered in all TSC patient with DRE and
consistent semiology, concordant with EEG and MRI findings. On the
other hand, children with multiple seizure types, multiple cortical tu­
bers, and multiple EEG abnormalities remain a challenge for presurgical
evaluation and likely postsurgical seizure freedom will not be achieved.
The panel proposed that the identification of ideal candidates for sur­
gery should be performed at expert centers specialized in both TSC
management and epilepsy surgery and emphasized that it should be
tailored (with ancillary investigations as high-resolution source imag­
ing, single photon emission computed tomography (SPECT), or invasive
N. Specchio et al. European Journal of Paediatric Neurology 47 (2023) 25–34

Table 3 as an emerging treatment for TSC associated seizures.

Unanswered questions as insights for future studies. In a recent RCT, metformin reduced seizure frequency compared to
Questions placebo in 21 patients. Ideally, these results would need further
confirmation with adequate statistical power [111].
- What is the rationale for selecting the optimal add-on therapy or combination
therapy if the seizures persist after the use of two appropriately chosen anti-seizure The relatively small number of patients included in this RCT hamper
medication? May we talk about first line and beyond? firm conclusions, and at present this panel cannot recommend the use of
- What is the optimal timing for pre-symptomatic treatment and for how many years metformin.
do we have to maintain pre-symptomatic treatment?
- Which is the optimal first add-on treatment option after vigabatrin failure in focal
seizures?
8. Challenges for the future and unmet needs
- When should mTOR inhibitors be introduced?
- What is the efficacy and safety (short and long term) of mTOR inhibitors as pre- Several questions still remain unanswered, providing hypotheses for
symptomatic therapy commenced in children below 2 years of age? future studies (see Table 3).
- Why is mTOR inhibition ineffective for seizure control in about 50% of patients? Are
there other mTOR-independent mechanisms involved or is this mainly related to the
already ongoing process of epileptogenesis, and what are the interactions with other 9. Conclusions
signaling pathways?
- Can we determine whether earlier use of mTOR inhibitors benefits Epileptogenesis in TSC is a chronic and dynamic process. Different
neurodevelopment in TSC?
convergent and divergent mechanisms are involved in the establishment
- Which are the patients specific presurgical characteristics related to better or worse
surgical outcome, and does early surgery improve neurodevelopmental outcome?
of a pathological “network” capable of generating seizures and co-
- What are the main barriers related to the underuse of epilepsy surgery? Is this occurring neurodevelopmental disorders. mTORC1-dependent and -in­
related to the complexity of epileptogenic networks? dependent mechanisms may occur in parallel and interact throughout,
therefore combinatorial treatment strategies to engage different targets
involved in the epileptogenic network have been hypothesized and
recording), to the individual patient.
referred to as “Network therapy” [112]. In TSC this is still a hypothesis,
and a careful selection of patients for any combination treatment is
7. Other treatment options
important. More data on its modifying effect is needed before combi­
nation therapy can be recommended in daily clinical care. In addition,
Two recent studies confirmed that both ketogenic diet therapies (KD)
there are important observations of spontaneous remission of epilepsy
and vagal nerve stimulation (VNS) could be of benefit in reducing the
with age that need to be acknowledged [113].
seizure frequency in individuals with the refractory epilepsy, who are
To identify the contribution of mTORC1 dependent and independent
not suitable for epilepsy surgery [99–102]. In a meta-analysis on the role
mechanisms to the clinical phenotype and to explain failures with tar­
of VNS in genetic epilepsies, VNS showed a significant effect on the
geted treatment with mTOR inhibitors, exploring combination treat­
proportion of patients with ≥50% seizure reduction (Z = 5.75, p <
ment strategies may be of interest. There is recent experimental
0.001). Heterogeneity (I2 = 40.96%, p = 0.06) and publication bias (p
evidence showing that ASD phenotypes in TSC may result from a com­
= 0.772) were non-significant. In the same meta-analysis the pooled
bination of TSC1/TSC2 molecular deficits and additional social im­
(95% CI) seizure freedom rate was 40% (12%–71%), ≥90% seizure
pairments caused by seizures, but through different mechanisms that
reduction rate was 31% (8%–56%), and ≥50% reduction rate was 68%
may not all respond to mTOR inhibitors [87].
(48%–91%) [100]. In a cost analysis of competing therapeutic options
Compared to the recommendations compiled 5 years ago, the panel
for drug resistant seizures associated with TSC, a probability of positive
believes that substantial progress has been made thanks to the early
outcome (Engel class 1) was 0.55 for resective surgery, 0.12 for VNS,
identification of risk factors, which can predict both the impending
0.00 for KD, and 0.20 for mTOR inhibition, respectively [103]. KD can
epilepsy and the possibility of a severe drug-resistant epilepsy.
be an effective therapeutic strategy for seizures particularly in infancy
Furthermore, pre-symptomatic assessment and treatment with vigaba­
and early childhood [101,102].
trin are now more and more adopted in very high-risk infants. The panel
In an international placebo controlled randomized clinical trial to
concluded that the clinical approach for TSC-related epilepsy has vastly
evaluate the efficacy and safety of cannabidiol (CBD) in 224 patients
improved over the last few years, based on the progress in predicting and
with TSC associated seizures, CBD at dosages of 25 or 50 mg/kg/day
modifying strategies and on the advent of molecular targeted therapy.
significantly reduced seizure frequency if compared with placebo, with
Progress in the knowledge of the mechanism underlying epileptogenesis
no significant difference in efficacy between 25 and 50 mg/kg/day. The
has paved the way for new concepts in the management for TSC-related
most common adverse events were diarrhea, somnolence and elevated
epilepsy. Developmental neurobiology and neuropathology offer a
transaminase, more often seen at higher dosages [20]. A good safety and
practical basis for approaching such concepts, and early genetic diag­
efficacy profile was reported in 199 patients who entered the open label
nosis and use of reliable EEG and MRI biomarkers may allow the
study to evaluate long-term safety and efficacy with add-on CBD [104].
development of pre-symptomatic and disease-modifying strategies. The
Importantly, CBD is known to significantly increase serum levels of
panel acknowledged the need to develop and validate a scoring method
mTOR inhibitors and such combined therapy requires dosing modifi­
to calculate the relative risk of impending epilepsy and timing of pre­
cation and close monitoring. In patients taking both CBD and ever­
ventive treatment. The goal is to develop a more robust and accurate
olimus, the everolimus dosage should be reduced by nearly a half, and
prediction model that importantly guides counseling and treatment.
through levels should be monitored regularly [105,106].
Based on the results of the EPISTOP trial, the panel emphasized the
CBD is hypothesized to act at a synaptic level by reduction in
importance of a close developmental follow-up of infants with TSC at a
excitatory neurotransmission through at least three possible mecha­
sensitive developmental timepoints (6, 9, 12 months) for an early
nisms: block of GPR55 receptors, desensitization of TRPV1 channels, or
detection of possible delay in motor, language or cognitive develop­
inhibition of ENT1 adenosine reuptake pumps [107,108]. Interestingly,
mental trajectories. Early intervention strategies are crucial to improve
CBD is also shown to be a selective modulator of the mTOR downstream
the neurodevelopmental outcome. All infants with a definite diagnosis
molecule rpS6 in the brain in a preclinical-model of TSC [109].
of TSC should be followed up with early and frequent EEG monitoring
Ganaxolone - a positive allosteric modulator of GABA a receptor –
(every 4 weeks in first 6–8 months and every 4–8 weeks thereafter until
was approved within the USA for the treatment of seizures associated
12 months and every 12 weeks between 12 and 24 months) to detect the
with CDKL5 deficiency disorder (CDD) in March 2022 [110], its use is
appearance of interictal epileptiform discharges and abnormal back­
under evaluation in a phase III study (TRUSTTSC) and could play a role
ground patterns. Pre-symptomatic treatment with vigabatrin as soon as

30
N. Specchio et al.
Table 4
Summary of the therapeutic recommendations for TSC associated seizures
(modified and updated from Curatolo P, Nabbout R, Lagae L et al. Management
of epilepsy associated with tuberous sclerosis complex: Updated clinical rec­
ommendations. Eur J Paediatr Neurol. 2018; 22 (5):738–748).
Treatment
Antiseizure medications
Vigabatrin
• Recommended as first-line mono­
therapy in TSC related spasms or focal
seizures in children <1 year of age
• Pre-symptomatic treatment with VGB
(see text for recommended time points
for clinical and EEG screening and
EEG indicators for initiating
treatment) is now strongly
recommended and may improve long-
term epilepsy and neurodevelopment
outcomes [18]
Other ASMs
• ACTH and prednisolone are effective
for TSC-related infantile spasms and
are used as second-line therapy
ASMs combination therapy
• Appropriate when Vigabatrin and
steroid therapy has failed
mTOR inhibitors: everolimus and sirolimus
• Everolimus is effective as an
adjunctive treatment in patients aged
2 years and older with refractory focal • Glucose intolerance and
seizures, with or without evolution to
focal to bilateral tonic-clonic seizures
• Everolimus should be considered
before the age of 6 years as in real-
world clinical practice better response
was seen in this age group
• Everolimus adjunctive therapy can be
considered after failure of 2
appropriate ASMs
• Sirolimus has a favorable safety and
efficacy profile for treating
manifestations of TSC (including
epilepsy)
Surgery
• Currently underutilized and under
assessed in TSC-associated refractory
seizures
• Presurgical evaluation should be
considered in all TSC patient with
DRE
• Early surgery significantly increases
probability that patients will be
seizure-free
• Resection beyond tuber margins is
associated with greater probability of
seizure freedom
• Multifocal and bilateral lesions do not
preclude presurgical assessment or
subsequent resective surgery
• Overall post-operative seizure
freedom is achieved in 50–60% of TSC
patients, and >50% seizure reduction
is seen in another 15%, even in pa­
tients with multiple tubers
Ketogenic diet
• Ketogenic dietary therapies can be an
effective therapeutic strategy for
seizures particularly in infancy and
early childhood
• Consider for patients who are not
surgical candidates, who have failed
surgery, or with multifocal seizure
onset
• Use instead of other ASMs after
vigabatrin failure remains
controversial
Limitations
• Retinal toxicity (lack of data in infants
and young children)
• Retinal toxicity concerns, newer data
suggest that this may be a lower risk
than previously thought [72]
• Careful selection for maximal synergy
and minimal unfavorable reactions/
toxicity
• Potential drug-drug interactions
• Seizures may recur when discontinued
immunosuppression.
• Children with multiple seizure types,
multiple cortical tubers, and multiple
EEG abnormalities remain a challenge
and likely postsurgical seizure
freedom will not be achieved but
palliative approach can yield clinical
improvement
• Compliance may be a problem in
children past infancy
31
European Journal of Paediatric Neurology 47 (2023) 25–34
Table 4 (continued )
Treatment Limitations
• Decreased mTOR activation in animal
models may provide biological basis
for mechanism of action in TSC-
associated seizures
Vagus nerve stimulation
• May be a first option in drug resistant • Almost no patients become seizure-
epilepsy when ketogenic diet is not free
acceptable or may be used in
combination with ketogenic diet
• Limited data indicate many patients
experience reduction of seizure
frequency
Cannabidiol
• CBD at the dosage of 25 up to 50 mg/ • Important drug-drug interaction with
kg/day significantly reduces seizure everolimus should be considered.
frequency if compared with placebo
• Dosages of 50 mg/kg/day are
associated with more adverse events
TSC: tuberos sclerosis complex, ACTH: Adrenocorticotropic hormone, mTOR:
mammalian target of rapamycin, ASM: anti-seizure medication.
focal spikes occur, but when multifocal spikes, pre-hypsarrhythmia/
hypsarrhythmia, subclinical seizures are recorded on EEG, delays sei­
zures and DRE risk, and should be used in all patients. The panel
emphasized the need to identify the subpopulation of infants that have
an even higher risk to develop epilepsy and ASD. Moreover, parents
should be trained to recognize the appearance of subtle focal seizures or
atypical behavior or development early. All infants with a definite
diagnosis of TSC should be evaluated with standard developmental
scales at 6, 12, 18 and 24 months to detect early deviations in motor,
language, and cognitive trajectories. Initial neurodevelopmental atypias
in children with TSC may represent a risk of later ASD diagnosis. A
summary of the therapeutic recommendations for TSC associated sei­
zures is given in Table 4.
The panel is pleased to acknowledge the great progress made in the
management of TSC- associated epilepsy and believes that a combina­
tion of different therapeutic options could be considered in a well
selected population of children, even if a personalized medicine
approach remains a challenge for future years.
Declaration of competing interest
We confirm that we have read the Journal’s position on issues
involved in ethical publication and affirm that this report is consistent
with those guidelines.
N Specchio has served on scientific advisory boards for GW Pharma,
BioMarin, Arvelle, Marinus and Takeda; has received speaker honoraria
from Eisai, Biomarin, Livanova, Sanofi; has served as an investigator for
Zogenix, Marinus, Biomarin, UCB, Roche.
R Nabbout has served as principal investigators in clinical trials for
Novartis, Nutricia, Eisai, UCB, GW Pharma, Livanova. She received
consulting and lecturer honoraria from Biogene, BioMarin, Praxis, GW
Pharma, Zogenix, Novartis, Nutricia, Stoke, Ionis, Targeon, Neu­
raxpharma, Takeda, Nutricia, Biocodex, Advicennes and Eisai. She
received unrestricted research grants from Eisai, UCB, Livanova and GW
Pharma and academic research grants from EJP-RD (horizons 2020).
E Aronica has nothing to disclose.
S Auvin is a deputy editor at Epilepsia and has received support from,
and/or has served as a paid consultant for Biocodex, Eisai, Encoded, Jazz
Pharmaceuticals, GRIN Therapeutics, Neuraxpharm, Nutricia, Orion
Pharma, Supernus Pharmaceuticals, Takeda Pharmaceutical Company,
UCB Pharma, Vitaflo, Xenon Pharmaceuticals, and Zogenix.
M Feucht received speaker and advisory board honoraria as well as
scientific grants from Biocodex, Eisai, Gerot, Livanova, Novartis,
Nutricia, Shire, UCB and GW Pharma.
F Jansen has served as a consultant for Jazz Pharmaceuticals,
N. Specchio et al.
Nutricia, Novartis and UCB. She was an investigator of the GWEPCARE
1512 trial.
L Lagae received grants and is a consultant and/or speaker for
Zogenix; LivaNova, UCB, Shire, Eisai, Novartis, Takeda/Ovid, NEL,
Epihunter.
A Benvenuto, K Kotulska, S Jozwiak, P Curatolo report no conflicts of
interest.
Acknowledgments
The contribution of Sergiusz Jozwiak and Katarzyna Kotulska has
been partly supported by grant EPIMARKER of the Polish National
Center for Research and Development no. STRATEGMED3/306306/4/
2016, grant VIRAP (Project number 2019/ABM/01/00034) and grant
RaRE-TS (Project number 2020/ABM/01/00054) of the Medical
Research Agency, Poland, and a statutory grant of the Children’s Me­
morial Health Institute financed by the Polish Ministry of Science and
Higher Education.
Eleonora Aronica acknowledges support by The Netherlands Orga­
nisation for Health Research and Development (ZonMw).
Rima Nabbout, Eleonora Aronica, Arianna Benvenuto, Martha
Feucht, Floor Jansen, Katarzyna Kotulska, Lieven Lagae, Sergiusz Joz­
wiak, and Paolo Curatolo were supported by the 7th Framework Pro­
gram of the European Commission within the Large-Scale Integrating
Project EPISTOP (Proposal No. 602391–2; www.epistop.eu).
Nicola Specchio is supported by #NEXTGENERATIONEU (NGEU)
and funded by the Ministry of University and Research (MUR), National
Recovery and Resilience Plan (NRRP), project MNESYS (PE0000006) –
A Multiscale integrated approach to the study of the nervous system in
health and disease (DN. 1553 October 11, 2022).
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