Inhibition of p70 Ribosomal S6 Kinase (S6K1) Reduces Cortical Blood Flow in a Rat Model of Autism-Tuberous Sclerosis.

The manifestations of tuberous sclerosis complex (TSC) in humans include epilepsy, autism spectrum disorders (ASD) and intellectual disability. Previous studies suggested the linkage of TSC to altered cerebral blood flow and metabolic dysfunction. We previously reported a significant elevation in cerebral blood flow in an animal model of TSC and autism of young Eker rats. Inhibition of the mammalian target of rapamycin (mTOR) by rapamycin could restore normal oxygen consumption and cerebral blood flow. In this study, we investigated whether inhibiting a component of the mTOR signaling pathway, p70 ribosomal S6 kinase (S6K1), would yield comparable effects. Control Long Evans and Eker rats were divided into vehicle and PF-4708671 (S6K1 inhibitor, 75 mg/kg for 1 h) treated groups. Cerebral regional blood flow (14C-iodoantipyrine) was determined in isoflurane anesthetized rats. We found significantly increased basal cortical (+ 32%) and hippocampal (+ 15%) blood flow in the Eker rats. PF-4708671 significantly lowered regional blood flow in the cortex and hippocampus of the Eker rats. PF-4708671 did not significantly lower blood flow in these regions in the control Long Evans rats. Phosphorylation of S6-Ser240/244 and Akt-Ser473 was moderately decreased in Eker rats but only the latter reached statistical significance upon PF-4708671 treatment. Our findings suggest that moderate inhibition of S6K1 with PF-4708671 helps to restore normal cortical blood flow in Eker rats and that this information might have therapeutic potential in tuberous sclerosis complex and autism.

Tuberous sclerosis complex in adulthood: focus on epilepsy prognosis.

OBJECTIVE: Typically diagnosed in early childhood or adolescence, TSC is a chronic, multisystemic disorder with age-dependent manifestations posing a challenge for transition and for specific surveillance throughout the lifetime. Data on the clinical features and severity of TSC in adults and on the prognosis of epilepsy are scarce. We analyzed the clinical and genetic features of a cohort of adult patients with TSC, to identify the prognostic predictors of seizure remission after a long follow-up. METHOD: We conducted a retrospective analysis of patients diagnosed with TSC according to the updated international diagnostic criteria. Pearson's chi-square or Fisher's exact test and Mann Whitney U test were used to compare variables among the Remission (R) and Non-Remission (NR) group. Univariate and multivariate logistic regression analyses were performed. RESULTS: We selected 43 patients with TSC and neurological involvement in terms of epilepsy and/or brain lesions, attending the Epilepsy Center of our Institute: of them, 16 (37.2%) were transitioning from the pediatric care and 6 (13.9%) were referred by other specialists. Multiorgan involvement includes cutaneous (86.0%), nephrological (70.7%), hepatic (40.0%), ocular (34.3%), pneumological (28.6%) and cardiac (26.3%) manifestations. Thirty-nine patients (90.7 %) had epilepsy. The mean age at seizure onset was 4 ± 7.3 years: most patients (29, 76.3 %) presented with focal seizures or spasms by age 3 years; only 2 (5.3 %) had seizure onset in adulthood. Twenty-seven patients (69.2 %) experienced multiple seizure types overtime, 23 (59.0 %) had intellectual disability (ID). At last assessment, 14 (35.9 %) were seizure free (R group) and 25 (64.1 %) had drug-resistant seizures (NR group). At logistic regression univariate analysis, ID (OR 7.9, 95 % CI 1.8--34.7), multiple seizure types lifelong (OR 13.2, 95 % CI 2.6- 67.2), spasms/tonic seizures at presentation (OR 6.5, 95 % CI 1.2--35.2), a higher seizure frequency at onset (OR 5.4, 95 % CI 1.2--24.3), abnormal neurological examination (OR 9.8, 95 % CI 1.1--90.6) and pathogenic variants in TSC2 (OR 5.4, 95 % CI 1.2--24.5) were significantly associated with non-remission. In the multivariate analysis, both ID and multiple seizure types lifelong were confirmed as independent predictors of poor seizure outcome. CONCLUSIONS: In our cohort of adult patients with TSC, epilepsy remains one of the main neurological challenges with only 5.3% of cases manifesting in adulthood. Approximately 64% of these patients failed to achieve seizure remission. ID and multiple seizure types were the main predictors of poor outcome. Nephrological manifestations require continuous specific follow-up in adults.

Treatment guidelines for rare, early-onset conditions associated with epileptic seizures: a literature review on Rett syndrome and tuberous sclerosis complex.

BACKGROUND: Rett syndrome (RTT) and tuberous sclerosis complex (TSC) are two rare disorders presenting with a range of different epileptic seizures. Seizure management requires careful therapy selection, thereby necessitating development of high-quality treatment guidelines. This targeted literature review (TLR) aimed to characterise country-specific and international treatment guidelines available for pharmacological management of seizures in RTT and TSC. METHODS: A TLR was performed between 25-Jan and 11-Mar 2021. Manual searches of online rare disease and guideline databases, and websites of national heath technology assessment bodies were conducted for the following countries: Australia, Canada, France, Germany, Israel, Italy, Japan, Spain, Switzerland, UK, and US as defined by pre-specified eligibility criteria. Search terms were developed for each condition and translated into local languages where appropriate. Eligible publications were defined as guidelines/guidance reporting pharmacological management of seizures in patients with RTT and TSC. Guideline development methodology, geographical focus, author information and treatment recommendations were extracted from guidelines. An author map was generated using R version 3.5.1 to visualise extent of collaboration between authors. RESULTS: 24 total guidelines were included, of which three and six contained only recommendations for RTT and TSC, respectively (some provided recommendations for ≥ 1 condition). Guideline development processes were poorly described (50% [12 guidelines] had unclear/absent literature review methodologies); reported methodologies were variable, including systematic literature reviews (SLRs)/TLRs and varying levels of expert consultation. Most (83% [20/24]) were country-specific, with guideline authors predominantly publishing in contained national groups; four guidelines were classified as 'International,' linking author groups in the US, UK, Italy and France. High levels of heterogeneity were observed in the availability of treatment recommendations across indications, with 13 and 67 recommendations found for RTT and TSC, respectively. For RTT, all treatment recommendations were positive and sodium valproate had the highest number of positive recommendations (Khwaja, Sahin (2011) Curr Opin Pediatr 23(6):633-9). All TSC treatments (21 medications) received either exclusively negative (National Organization for Rare Disorders (2019)) or positive (Chu-Shore et al. (2010) Epilepsia 51(7):1236-41) recommendations; vigabatrin received the highest number of positive recommendations (Kaur, Christodoulou (2019)). CONCLUSIONS: This review highlights the need for the development of international high-quality and comprehensive consensus-based guidance for the management of seizures with pharmacological therapy in RTT and TSC. TRIAL REGISTRATION: Not applicable.

Cannabidiol (Epidyolex®) for severe behavioral manifestations in patients with tuberous sclerosis complex, mucopolysaccharidosis type III and fragile X syndrome: protocol for a series of randomized, placebo-controlled N-of-1 trials.

BACKGROUND: Many rare genetic neurodevelopmental disorders (RGNDs) are characterized by intellectual disability (ID), severe cognitive and behavioral impairments, potentially diagnosed as a comorbid autism spectrum disorder or attention-deficit hyperactivity disorder. Quality of life is often impaired due to irritability, aggression and self-injurious behavior, generally refractory to standard therapies. There are indications from previous (case) studies and patient reporting that cannabidiol (CBD) may be an effective treatment for severe behavioral manifestations in RGNDs. However, clear evidence is lacking and interventional research is challenging due to the rarity as well as the heterogeneity within and between disease groups and interindividual differences in treatment response. Our objective is to examine the effectiveness of CBD on severe behavioral manifestations in three RGNDs, including Tuberous Sclerosis Complex (TSC), mucopolysaccharidosis type III (MPS III), and Fragile X syndrome (FXS), using an innovative trial design. METHODS: We aim to conduct placebo-controlled, double-blind, block-randomized, multiple crossover N-of-1 studies with oral CBD (twice daily) in 30 patients (aged ≥ 6 years) with confirmed TSC, MPS III or FXS and severe behavioral manifestations. The treatment is oral CBD up to a maximum of 25 mg/kg/day, twice daily. The primary outcome measure is the subscale irritability of the Aberrant Behavior Checklist. Secondary outcome measures include (personalized) patient-reported outcome measures with regard to behavioral and psychiatric outcomes, disease-specific outcome measures, parental stress, seizure frequency, and adverse effects of CBD. Questionnaires will be completed and study medication will be taken at the participants' natural setting. Individual treatment effects will be determined based on summary statistics. A mixed model analysis will be applied for analyzing the effectiveness of the intervention per disorder and across disorders combining data from the individual N-of-1 trials. DISCUSSION: These N-of-1 trials address an unmet medical need and will provide information on the effectiveness of CBD for severe behavioral manifestations in RGNDs, potentially generating generalizable knowledge at an individual-, disorder- and RGND population level. TRIAL REGISTRATION: EudraCT: 2021-003250-23, registered 25 August 2022, https://www.clinicaltrialsregister.eu/ctr-search/trial/2021-003250-23/NL .

The association between computed tomography attenuation value of renal angiomyolipoma associated with tuberous sclerosis complex and response to everolimus.

BACKGROUND: The response to everolimus in patients with renal angiomyolipoma associated with tuberous sclerosis complex (TSC-RAML) varies among individuals. This study aims to identify potential factors associated with the response to everolimus. METHOD: We retrospectively examined data encompassing age, gender, tumor size, computed tomography attenuation value (CT value), CT enhancement, and tumor reduction rate in patients with TSC-RAML undergoing everolimus in two previously registered clinical trials. RESULT: A total of 33 participants (29.33 ± 6.63 years old, 20 females) were included. The correlation analysis conducted separately for tumors located in the left and right kidneys revealed significant negative correlations (P < 0.05) between tumor reduction rate and age, as well as tumor size. While significant positive correlations (P < 0.05) were observed between tumor reduction rate and unenhanced CT value as well as CT enhancement. Nonetheless, based on multiple linear regression analysis, unenhanced CT value emerged as the sole-independent predictor of tumor reduction rate among age, gender, tumor size, unenhanced CT value and CT enhancement for both left (coefficient = 0.00319, P < 0.0001) and right kidneys (coefficient = 0.00315, P = 0.0104). Notable reductions were observed in unenhanced CT value (- 3.81 vs - 24.70HU, P < 0.0001) and CT enhancement (48.16 vs 33.56HU, P < 0.0001) following a 3-month administration of everolimus. The decline in both unenhanced CT value and tumor size predominantly occurred within the initial 3 months, subsequently maintaining a relatively stable level throughout the treatment. CONCLUSION: The unenhanced CT value of TSC-RAML showed an independent correlation with the response to everolimus, suggesting its potential as a predictor of everolimus efficacy in patients with TSC-RAML.

An updated meta-analysis of effectiveness and safety of mTOR inhibitors in the management of tuberous sclerosis complex patients.

PURPOSE: Tuberculous sclerosis complex (TSC) is an autosomal dominant multi-system disease. In TSC patients, the inhibition of mTOR pathway is weakened, which leads to the uncontrolled proliferation of normal resting cells. Therefore, mTOR inhibitors have many therapeutic potentials in the treatment of TSC. However, there is no consensus on the safety and efficacy of mTOR inhibitors so far. This article aimed to present new evidence for the efficacy and safety of mTOR inhibitors in the treatment of TSC by evaluating published clinical trials. METHODS: A systemic search of online databases, such as Cochrane Library, Embase, PubMed, and the US National Institutes of Health Clinical Trials Registry, was conducted. The researchers selected studies that met the following entry criteria: randomized, double-blinded or single-blinded, placebo-controlled, parallel-group studies with active and control arms receiving rapamycin or everolimus and matched placebo, respectively. The meta-analysis included seven studies. Tumor response or epilepsy seizure frequency response rates were considered efficacy outcomes. RESULTS: In seven studies involving 877 patients, using of mTOR inhibitors therapy showed an improvement in both tumor response and seizure frequency outcomes in TSC. In combination of AML (angiomyolipomas), SEGA (subependymal giant cell astrocytoma), epilepsy, and facial angiofibroma subjects, the RR is 3.01 (95% CI 2.03 to 4.45, p = 0.000) with observed heterogeneity (I-squared = 55.4%). The main side effect of mTOR inhibitors was stomatitis. CONCLUSION: The updated meta-analysis suggests that the use of mTOR inhibitors is an effective therapy for patients with TSC.

Treatment of tuberous sclerosis complex manifestations in children with mTOR inhibitors.

PURPOSE: Tuberous sclerosis complex (TSC) is a rare autosomal dominant genetic disorder that affects multiple organ systems. Mutations in the TSC1 and TSC2 genes result in the constitutive hyperactivation of the mammalian target of rapamycin (mTOR) pathway, contributing to the growth of benign tumors or hamartomas in various organs. Due to the implication of mTOR pathway dysregulation in the disease pathology, increasing evidence supports the use of mTOR inhibitors for treating multiple manifestations of TSC. METHODS: In this study, we conducted a retrospective analysis of clinical findings and treatment data from 38 patients diagnosed with tuberous sclerosis who were followed up in the Pediatric Oncology Clinic between 2010 and 2020. We collected information on patients' ages, genders, affected sites, familial history, imaging findings, presence of tumors, and treatments. RESULTS: Among the patients, nine individuals with TSC manifestations were treated with mTOR inhibitors. Specifically, everolimus was successfully administered to five patients with inborn cardiac rhabdomyoma causing hemodynamic impairment. In addition, two patients with refractory seizures received everolimus in combination with anti-epileptic drugs. A patient with renal angiomyolipomas larger than 3 cm was treated with everolimus, while a patient with extensive facial angiofibroma received topical sirolimus. All patients tolerated the mTOR inhibitors well, and the side effects were deemed acceptable. CONCLUSION: The utilization of mTOR inhibition in TSC is expected to become more prevalent in clinical practice, as current research is anticipated to provide a better understanding of the therapeutic roles of these treatments in TSC.

Complete Remission of a Diffuse Large B-Cell Lymphoma in a Young Patient, with Severe Tuberous Sclerosis, Treated with Metronomic Chemotherapy and Ibrutinib: A Case Report.

Tuberous sclerosis (TS) is a rare autosomal dominant genetic multisystem disease caused by mutations in either the TSC1 or TSC2 gene and results in the growth of non-cancerous masses in several organs. Diffuse large B-cell lymphoma (DLBCL) is the predominant non-Hodgkin lymphoma in adolescents and young adults. Metronomic chemotherapy (mCHEMO) can be defined as the frequent, regular administration of drug doses able to maintain a low, but active, range of concentrations of chemotherapeutic drugs during prolonged periods of time. We present the case of a young woman with severe TS who developed DLBCL. She was treated consecutively with the mCHEMO schedule R-DEVEC (prednisone, vinorelbine, etoposide, cyclophosphamide, plus rituximab) and then ibrutinib, achieving an impressive long-lasting complete remission. In conclusion, alternative treatments could be necessary when comorbidities are present in patients, and mCHEMO can be a potential successful therapeutic approach in frail subjects.

Long-term use of everolimus for refractory arrhythmia in a child with tuberous sclerosis complex.

Tuberous sclerosis complex is associated with the occurrence of cardiac rhabdomyomas that may result in life-threatening arrhythmia unresponsive to standard antiarrhythmic therapy. We report the case of an infant with multiple cardiac rhabdomyomas who developed severe refractory supraventricular tachycardia (SVT) that was successfully treated with everolimus. Pharmacological mTOR inhibition rapidly improved arrhythmia within few weeks after treatment initiation and correlated with a reduction in tumor size. Intermediate attempts to discontinue everolimus resulted in rhabdomyoma size rebound and recurrence of arrhythmic episodes, which resolved on resumption of therapy. While everolimus treatment led to successful control of arrhythmia in the first years of life, episodes of SVT reoccurred at the age of 6 years. Electrophysiologic testing confirmed an accessory pathway that was successfully ablated, resulting in freedom of arrhythmic events. In summary we present an in-depth evaluation of the long-term use of everolimus in a child with TSC-associated SVT, including the correlation between drug use and arrhythmia outcome. This case report provides important information on the safety and efficacy of an mTOR inhibitor for the treatment of a potentially life-threatening cardiac disease manifestation in TSC for which the optimal treatment strategy is still not well established.

Effectiveness and safety of everolimus treatment in patients with tuberous sclerosis complex in real-world clinical practice.

BACKGROUND: The randomised double-blinded placebo-controlled EXIST-1-3 studies have showed everolimus effective with adverse effects reported as acceptable in treatment of symptoms in patients with tuberous sclerosis complex (TSC), although evidence of outcomes in clinical practice remains limited. This study aimed to investigate, in clinical practice, the effectiveness and safety of everolimus for epilepsy, renal angiomyolipoma (rAML), and subependymal giant cell astrocytoma (SEGA) in patients with TSC. RESULTS: The study included 64 patients with TSC (median age: 19, range 0.9-54 years) receiving everolimus treatment (Norway: n = 35; Denmark: n = 29). Among 45 patients with epilepsy, 14 (31%) were responders experiencing ≥ 50% reduction in seizure frequency in the last 3 months of treatment compared with the last 3 months before treatment. Nineteen (42%) patients changed their anti-seizure medications (ASMs). Responders were more common among patients < 18 years (46%) than among patients ≥ 18 years (14%, p = 0.03). In 29 patients with rAML, everolimus reduced (≥ 30% decrease) and stabilized (< 20% increase, ≤ 30% decrease) longest diameter of rAML in 38% and 59%, respectively, after a mean treatment duration of 37 months. SEGA volume was reduced in three patients by 71%, 43%, and 48% after 39, 34, and 82 months. Adverse effects were reported in 61 of 64 patients (95%) after a median treatment duration of 31 months (range 0-106), with oral ulceration/stomatitis (63%) and upper respiratory tract infections (38%) being the most common. The most common laboratory abnormalities were increased cholesterol (41%), anaemia (30%), and leucopoenia (25%). Grade 3-4 adverse effects were reported in 36% of cases, and life-threatening conditions were reported in two patients. Nine patients discontinued everolimus treatment. CONCLUSIONS: Seizure reduction in this study sample was consistent with results from EXIST, but might be lower than expected, given that changes in concomitant ASMs are part of clinical practice. Seizure reduction was associated with younger age. As with EXIST, everolimus reduced or stabilised rAML size in most patients. SEGA volume was reduced in all three patients. Close follow-up is needed for this group, especially for children and patients who may not be able to report adverse effects.

Predicting Antiseizure Medication Treatment in Children with Rare Tuberous Sclerosis Complex-Related Epilepsy Using Deep Learning.

BACKGROUND AND PURPOSE: Tuberous sclerosis complex disease is a rare, multisystem genetic disease, but appropriate drug treatment allows many pediatric patients to have positive outcomes. The purpose of this study was to predict the effectiveness of antiseizure medication treatment in children with tuberous sclerosis complex-related epilepsy. MATERIALS AND METHODS: We conducted a retrospective study involving 300 children with tuberous sclerosis complex-related epilepsy. The study included the analysis of clinical data and T2WI and FLAIR images. The clinical data consisted of sex, age of onset, age at imaging, infantile spasms, and antiseizure medication numbers. To forecast antiseizure medication treatment, we developed a multitechnique deep learning method called WAE-Net. This method used multicontrast MR imaging and clinical data. The T2WI and FLAIR images were combined as FLAIR3 to enhance the contrast between tuberous sclerosis complex lesions and normal brain tissues. We trained a clinical data-based model using a fully connected network with the above-mentioned variables. After that, a weighted-average ensemble network built from the ResNet3D architecture was created as the final model. RESULTS: The experiments had shown that age of onset, age at imaging, infantile spasms, and antiseizure medication numbers were significantly different between the 2 drug-treatment outcomes (P < .05). The hybrid technique of FLAIR3 could accurately localize tuberous sclerosis complex lesions, and the proposed method achieved the best performance (area under the curve = 0.908 and accuracy of 0.847) in the testing cohort among the compared methods. CONCLUSIONS: The proposed method could predict antiseizure medication treatment of children with rare tuberous sclerosis complex-related epilepsy and could be a strong baseline for future studies.

Retrospective pharmacogenetic study in a cohort of pediatric tuberous sclerosis complex patients using everolimus.

Aim: Tuberous sclerosis complex (TSC) is a rare disease that produces multisystemic disorders. Everolimus (EVR) is the only immunosuppressive drug approved to control the symptoms and progression of the disease. The aim was to evaluate the genotype-phenotype association to improve the pediatric TSC pharmacotherapeutic outcome. Patients & methods: Ten pediatric TSC patients were recruited. Concomitant treatment and main metabolic enzymes and transporter coding gene variants of EVR were analyzed. Results: Significant associations were found between CYP3A4*22 allele and concomitant treatment with valproic acid (CYP3A4-inhibitor) with a poor metabolizer phenotype and the presence of pneumonia. Conclusion: This is the first pharmacogenetic study of EVR in pediatric TSC patients. The authors propose to consider concomitant treatment and pharmacogenetics due to their multifactorial status.

Adverse Events of Everolimus in Patients with Tuberous Sclerosis Complex Treated for Renal Angiomyolipoma/Subependymal Giant Cell Astrocytoma.

Background: Although regarded as a potentially efficient approach to address tuberous sclerosis complex (TSC)-associated complications, the adverse event profile of everolimus has not yet been fully elucidated. The present study aimed to clarify the adverse event spectrum in patients with TSC who are using everolimus for common indications, in comparison to those who do not use everolimus. Materials and Methods: We recruited patients with TSC who were followed up annually at TSC integrated clinics or referred for medical assistance. Medical reviews and laboratory investigations were performed at baseline and annually by clinical physicians. The adverse events were assessed as per the National Cancer Institute Common Terminology Criteria for Adverse Events. Results: Common adverse events in everolimus users included hypercholesterolemia (55%), gingivostomatitis (50%), proteinuria (50%), and hyperglycemia (40%). Compared with everolimus nonusers, the occurrence of gingivostomatitis and proteinuria was significantly higher in everolimus users (gingivostomatitis, p=0.02; proteinuria, p=0.02). Among the everolimus users, 12 patients had level I CTCAE, and five had level II CTCAE. None of the everolimus users presented with CTCAE level III or higher. Conclusion: Patients with TSC who are everolimus users had a higher tendency to develop gingivostomatitis and proteinuria compared to nonusers. However, no differences were observed in the occurrence of other adverse events between everolimus users and nonusers.

Real-world evidence on the use of cannabidiol for the treatment of drug resistant epilepsy not related to Lennox-Gastaut syndrome, Dravet syndrome or Tuberous Sclerosis Complex.

INTRODUCTION: Highly purified cannabidiol (CBD) has a broad spectrum of action and could be useful for the treatment of drug resistant epilepsy regardless of etiology or syndrome. MATERIALS AND METHODS: Multicenter retrospective study that evaluated the efficacy and safety of CBD for the treatment of drug resistant epilepsy of different etiologies in patients >2 years of age. RESULTS: Seventy-eight patients with a median age of 24 years and a wide spectrum of mainly structural and genetic etiologies were included. Patients were using a median of 3 antiseizure drugs (IQR=2-4) and had a median of 30 monthly seizures (IQR=12-100) before starting CBD. The median treatment time with CBD was 14 months (IQR=10-17). The efficacy analysis at the last available visit showed that mean percent reduction in seizures, ≥50% reduction in seizure frequency and seizure freedom was 67.8%, 68.8% and 11.5% respectively. We found no significant impact of concomitant clobazam use on the efficacy and safety of CBD. In the safety analysis, 28.2% (n = 22) of patients presented adverse events related to CBD and drug-retention rate was 78.2%. CONCLUSIONS: In a real-world setting, highly purified CBD has been shown to be safe and effective for the treatment of drug resistant epilepsy not related to Lennox-Gastaut syndrome, Dravet syndrome or Tuberous Sclerosis Complex. Based on these findings, highly purified CBD should be considered as an adjuvant therapy for drug resistant epilepsy, regardless of its underlying cause or specific syndrome. Nevertheless, this assumption should be validated through further controlled trials.

Paradigm shift in the treatment of tuberous sclerosis: Effectiveness of everolimus.

Tuberous sclerosis complex (TSC) is an autosomal dominant disease characterised by abnormal cell proliferation and differentiation that affects multiple organs and can lead to the growth of hamartomas. Tuberous sclerosis complex is caused by the disinhibition of the protein mTOR (mammalian target of rapamycin). In the past, various therapeutic approaches, even if only symptomatic, have been attempted to improve the clinical effects of this disease. While all of these therapeutic strategies are useful and are still used and indicated, they are symptomatic therapies based on the individual symptoms of the disease and therefore not fully effective in modifying long-term outcomes. A new therapeutic approach is the introduction of allosteric inhibitors of mTORC1, which allow restoration of metabolic homeostasis in mutant cells, potentially eliminating most of the clinical manifestations associated with Tuberous sclerosis complex. Everolimus, a mammalian target of the rapamycin inhibitor, is able to reduce hamartomas, correcting the specific molecular defect that causes Tuberous sclerosis complex. In this review, we report the findings from the literature on the use of everolimus as an effective and safe drug in the treatment of TSC manifestations affecting various organs, from the central nervous system to the heart.

Microscopic Kidney Disease in Tuberous Sclerosis Complex and Treatment With mTOR Inhibition.

Declining kidney function in tuberous sclerosis complex (TSC) is often attributed to large lesions, including angiomyolipomas (AMLs) and cysts, that encroach on the normal parenchyma or that require intervention and loss of parenchyma from surgical debulking or embolization. Consequently, research on inhibitors of the mammalian target of rapamycin (mTOR), a protein complex implicated in TSC pathophysiology for its role in promoting cell growth and proliferation, has largely focused on their ability to reduce AML size. Clinical guidelines distilled from this research limit mTOR inhibition as a first-line treatment to patients with large AMLs. However, chronic kidney disease (CKD) occurs in patients without large AMLs or a history of renal intervention. Alternate mechanisms postulated for CKD in TSC may suggest a role for mTOR inhibition in this population. In this report, we present 2 cases of a microscopic variant of TSC kidney disease causing declining kidney function, as well as anecdotal evidence for the use of mTOR inhibition to improve kidney function in the absence of large AMLs. We highlight the importance of annual kidney function assessment in patients with TSC and suggest a low threshold for kidney biopsy in patients with declining glomerular filtration rate without a clear etiology clinically or radiographically.

Seizure reduction in TSC2-mutant mouse model by an mTOR catalytic inhibitor.

OBJECTIVE: Tuberous sclerosis complex (TSC) is a neurodevelopmental disorder caused by autosomal-dominant pathogenic variants in either the TSC1 or TSC2 gene, and it is characterized by hamartomas in multiple organs, such as skin, kidney, lung, and brain. These changes can result in epilepsy, learning disabilities, and behavioral complications, among others. The mechanistic link between TSC and the mechanistic target of the rapamycin (mTOR) pathway is well established, thus mTOR inhibitors can potentially be used to treat the clinical manifestations of the disorder, including epilepsy. METHODS: In this study, we tested the efficacy of a novel mTOR catalytic inhibitor (here named Tool Compound 1 or TC1) previously reported to be more brain-penetrant compared with other mTOR inhibitors. Using a well-characterized hypomorphic Tsc2 mouse model, which displays a translationally relevant seizure phenotype, we tested the efficacy of TC1. RESULTS: Our results show that chronic treatment with this novel mTOR catalytic inhibitor (TC1), which affects both the mTORC1 and mTORC2 signaling complexes, reduces seizure burden, and extends the survival of Tsc2 hypomorphic mice, restoring species typical weight gain over development. INTERPRETATION: Novel mTOR catalytic inhibitor TC1 exhibits a promising therapeutic option in the treatment of TSC.

Cardiac rhabdomyomas: clinical progression, efficacy and safety of everolimus treatment.

BACKGROUND: Primary cardiac tumors are extremely rare. Cardiac rhabdomyoma is the most common primary cardiac tumor. 50-80% of solitary rhabdomyomas and all multiple rhabdomyomas are associated with tuberous sclerosis complex. Due to spontaneous regression, surgery is necessary only in severe hemodynamic compromise and persistent arrhythmias. Everolimus, a mechanistic target of rapamycin (mTOR) inhibitor, can be used in the treatment of rhabdomyomas seen in tuberous sclerosis complex. We aimed to evaluate the clinical progression of rhabdomyomas followed-up in our center between the years 2014-2019 and evaluate the efficacy and safety of everolimus treatment on tumor regression. METHODS: Clinical features, prenatal diagnosis, clinical findings, tuberous sclerosis complex presence, treatment and follow-up results were evaluated retrospectively. RESULTS: Among 56 children with primary cardiac tumors, 47 were diagnosed as rhabdomyomas, 28/47 patients (59.6%) had prenatal diagnosis, 85.1% were diagnosed before one year of age and 42/47 patients (89.3%) were asymptomatic. Multiple rhabdomyomas were present in 51% and median diameter of tumors was 16mm (4.5 - 52 mm). In 29/47 patients (61.7%) no medical or surgical treatment were necessary while 34% of these had spontaneous regression. Surgery was necessary in 6/47 patients (12.7%). Everolimus was used in 14/47 patients (29.8%). Indications were seizures (2 patients) and cardiac dysfunction (12 patients). Regression in size of rhabdomyomas was achieved in 10/12 patients (83%). Although, in the long-term, the amount of tumor mass shrinkage was not significantly different between patients who received everolimus and untreated patients (p=0.139), the rate of mass reduction was 12.4 times higher in patients who received everolimus. Leukopenia was not detected in any of the patients, but, hyperlipidemia was noted in 3/14 patients (21.4%). CONCLUSIONS: According to our results, everolimus accelerates tumor mass reduction, but not amount of mass regression in the long term. Everolimus may be considered for treatment of rhabdomyomas which cause hemodynamic compromise or life-threatening arrhythmias before surgical intervention.

Prdx5 in the Regulation of Tuberous Sclerosis Complex Mutation-Induced Signaling Mechanisms.

(1) Background: Tuberous sclerosis complex (TSC) mutations directly affect mTORC activity and, as a result, protein synthesis. In several cancer types, TSC mutation is part of the driver mutation panel. TSC mutations have been associated with mitochondrial dysfunction, tolerance to reactive oxygen species due to increased thioredoxin reductase (TrxR) enzyme activity, tolerance to endoplasmic reticulum (ER) stress, and apoptosis. The FDA-approved drug rapamycin is frequently used in clinical applications to inhibit protein synthesis in cancers. Recently, TrxR inhibitor auranofin has also been involved in clinical trials to investigate the anticancer efficacy of the combination treatment with rapamycin. We aimed to investigate the molecular background of the efficacy of such drug combinations in treating neoplasia modulated by TSC mutations. (2) Methods: TSC2 mutant and TSC2 wild-type (WT) cell lines were exposed to rapamycin and auranofin in either mono- or combination treatment. Mitochondrial membrane potential, TrxR enzyme activity, stress protein array, mRNA and protein levels were investigated via cell proliferation assay, electron microscopy, etc. (3) Results: Auranofin and rapamycin normalized mitochondrial membrane potential and reduced proliferation capacity of TSC2 mutant cells. Database analysis identified peroxiredoxin 5 (Prdx5) as the joint target of auranofin and rapamycin. The auranofin and the combination of the two drugs reduced Prdx5 levels. The combination treatment increased the expression of heat shock protein 70, a cellular ER stress marker. (4) Conclusions: After extensive analyses, Prdx5 was identified as a shared target of the two drugs. The decreased Prdx5 protein level and the inhibition of both TrxR and mTOR by rapamycin and auranofin in the combination treatment made ER stress-induced cell death possible in TSC2 mutant cells.