The Genetics of Tuberous Sclerosis Complex and Related mTORopathies: Current Understanding and Future Directions.

The mechanistic target of rapamycin (mTOR) pathway serves as a master regulator of cell growth, proliferation, and survival. Upregulation of the mTOR pathway has been shown to cause malformations of cortical development, medically refractory epilepsies, and neurodevelopmental disorders, collectively described as mTORopathies. Tuberous sclerosis complex (TSC) serves as the prototypical mTORopathy. Characterized by the development of benign tumors in multiple organs, pathogenic variants in TSC1 or TSC2 disrupt the TSC protein complex, a negative regulator of the mTOR pathway. Variants in critical domains of the TSC complex, especially in the catalytic TSC2 subunit, correlate with increased disease severity. Variants in less crucial exons and non-coding regions, as well as those undetectable with conventional testing, may lead to milder phenotypes. Despite the assumption of complete penetrance, expressivity varies within families, and certain variants delay disease onset with milder neurological effects. Understanding these genotype-phenotype correlations is crucial for effective clinical management. Notably, 15% of patients have no mutation identified by conventional genetic testing, with the majority of cases postulated to be caused by somatic TSC1/TSC2 variants which present complex diagnostic challenges. Advancements in genetic testing, prenatal screening, and precision medicine hold promise for changing the diagnostic and treatment paradigm for TSC and related mTORopathies. Herein, we explore the genetic and molecular mechanisms of TSC and other mTORopathies, emphasizing contemporary genetic methods in understanding and diagnosing the condition.

A large deletion in TSC2 causes tuberous sclerosis complex by dysregulating PI3K/AKT/mTOR signaling pathway.

BACKGROUND/AIM: Tuberous sclerosis complex (TSC) is a multi-system syndrome caused by loss-of-function mutation in TSC1 or TSC2. Most TSC patients present with cardiac rhabdomyoma or cortical tubers during fetal life, and the symptoms are not uniform as their age. The gene products of TSC1/2 are components of the TSC protein complex and are important role in the PI3K/AKT/mTOR (PAM) signaling pathway. Based on three members of a family with variable expressivity, the purpose of this study was to clarify the clinical features of TSC in different age groups and to analyze the genetic characteristics of TSC2 gene. METHODS: Clinical exome sequencing and co-segregation were used to identify a three-generation family with four affected individuals. HEK-293T cell model was constructed for subsequent experiments. Quantitative RT-PCR, western blotting, and subcellular localization were used to analyze the expression effect of TSC2 mutation. CCK-8 assay, wound healing assay, and cell cycle analysis were used to analyze the function effect of TSC2 mutation. RESULT: We identified a TSC family with heterozygous deletion of exon 4 in TSC2 by clinical exon sequencing. Sanger sequencing indicated that the affected individuals have 2541-bp deletion that encompassed exon 4 and adjacent introns. Deletion of exon 4 decreased the TSC2 mRNA and protein levels in HEK-293T cells, and activated the PI3K/AKT/mTOR pathway, thereby altering the cell cycle and promoting cell proliferation and migration. CONCLUSION: We confirmed the pathogenicity of the large deletion in TSC2 in a three- generations family.. Deletion of exon 4 of TSC2 affected cell proliferation, migration, and cell cycle via abnormal activation of the PAM pathway. This study evaluated the pathogenic effect of deletion of exon 4 of TSC2 and investigated the underlying mechanism.

Identification of a complex intrachromosomal inverted insertion in the long arm of chromosome 9 as a cause of tuberous sclerosis complex in a Korean family.

BACKGROUND: Tuberous sclerosis complex (TSC) is an autosomal dominant multisystem disorder, caused by a loss-of-function of either TSC1 or TSC2 gene. However, in 10%-15% TSC patients there is no pathogenic variant identified in either TSC1 or TSC2 genes based on standard clinical testing. METHODS: In this study, genome sequencing was performed for families with clinical diagnosis of TSC with negative results from TSC1 and TSC2 single-gene tests. RESULTS: Herein, we report a family presenting a classical TSC phenotype with an unusual, complex structural variant involving the TSC1 gene: an intrachromosomal inverted insertion in the long arm of chromosome 9. We speculate that the inverted 9q33.3q34.13 region was inserted into the q31.2 region with the 3'-end of the breakpoint of the inversion being located within the TSC1 gene, resulting in premature termination of TSC1. CONCLUSIONS: In this study, we demonstrate the utility of genome sequencing for the identification of complex chromosomal rearrangement. Because the breakpoints are located within the deep intronic/intergenic region, this copy-neutral variant was missed by the TSC1 and TSC2 single-gene tests and contributed to an unknown etiology. Together, this finding suggests that complex structural variants may be underestimated causes for the etiology of TSC.

Renal Cell Carcinoma Associated With TSC/MTOR Genomic Alterations: An Update on its Expanding Spectrum and an Approach to Clinicopathologic Work-up.

Renal cell carcinoma (RCC) with tuberous sclerosis complex (TSC)/mammalian target of rapamycin (MTOR) pathway-related genomic alterations have been classically described in hereditary TSC syndrome setting involving germline mutations, whereby cells with a bi-allelic inactivation of genes originate tumors in a classic tumor-suppressor "two-hit" Knudson paradigm. Initial studies of TSC-associated RCC categorized tumors into 3 broad heterogeneous morphologic groups: RCC with smooth muscle stroma, chromophobe-like, and eosinophilic-macrocytic. Recently, a similar morphologic spectrum has been increasingly recognized in novel and emerging entities characterized by somatic mutations in the TSC1/2 and MTOR in patients who do not suffer from the TSC. Correct recognition of RCC with TSC / MTOR mutations is critical for accurate prognostication because such tumors with aggressive behavior have the potential to be tailored to mTOR inhibitors. Whether TSC/MTOR mutated renal epithelial neoplasms represent a distinct molecular class has been confounded by the fact that TSC1/2 , and the gene encoding the downstream protein MTOR, are mutated secondarily in ∼5% of the more common subtypes of RCC, including the commonest subtype of clear cell RCC. This review summarizes the expanding morphologic spectrum of renal tumors with TSC/mTOR pathway alterations, specifically for sporadically occurring tumors where these genomic alterations likely are primary pathologic events. Finally, a practical surgical pathology approach to handling these tumors, and a conceptual framework of renal epithelial tumors with TSC/MTOR mutations as a "family of tumors", is presented.

Identification of a Novel TSC2 c.170G>A Missense Variant: A Case Report and Elaboration on the Yield of Targeted Options against Tuberous Sclerosis Complex Manifestations.

BACKGROUND: Tuberous sclerosis complex (TSC) is a rare genetic disease that affects multiple organs and affects the quality of life. Mutations in TSC1 and TSC2 genes are causing dysregulations in the mammalian target of the rapamycin (mTOR) pathway, inducing mostly benign but also malignant tumors, including renal cell carcinoma (RCC). The diagnosis of TSC, based on established clinical and genetic criteria, is essential for the optimal surveillance and management of patients. CASE PRESENTATION: With the current report, we present the case of two sisters who were consequently diagnosed with early-stage chromophobe-like RCC, possibly familial given their young age. The younger sister also had a previous diagnosis of differentiated thyroid carcinoma, for which she had been treated properly. Genetic testing of both revealed the same heterozygous TSC2 variant that is currently regarded as a variant of unknown significance, while both patients did not fulfill the clinical criteria for the diagnosis of TSC. Owing to these data, we opted to manage and surveil both sisters as TSC patients, while we also considered the specific TSC2 variant to be pathogenic - but of low penetrance - based on clinical judgment and functional analyses. Furthermore, we discussed the implementation of mTOR inhibitors for the treatment of TSC complications. CONCLUSION: As novel pathogenic variants of TSC genes are constantly being explored, the identification of TSC variants of unknown significance in combination with absent clinical diagnostic criteria cannot exclude a TSC diagnosis. We support the implementation of clinical judgment in assisting the diagnosis of TSC, as well as the enrollment of patients in clinical trials due to the rarity of the disease.

Non-canonical functions of a mutant TSC2 protein in mitotic division.

Tuberous Sclerosis Complex (TSC) is a debilitating developmental disorder characterized by a variety of clinical manifestations. TSC is caused by mutations in the TSC1 or TSC2 genes, which encode the hamartin/tuberin proteins respectively. These proteins function as a heterodimer that negatively regulates the mechanistic Target of Rapamycin Complex 1 (mTORC1). TSC research has focused on the effects of mTORC1, a critical signaling hub, on regulation of diverse cell processes including metabolism, cell growth, translation, and neurogenesis. However, non-canonical functions of TSC2 are not well studied, and the potential disease-relevant biological mechanisms of mutations affecting these functions are not well understood. We observed aberrant multipolar mitotic division, a novel phenotype, in TSC2 mutant iPSCs. The multipolar phenotype is not meaningfully affected by treatment with the inhibitor rapamycin. We further observed dominant negative activity of the mutant form of TSC2 in producing the multipolar division phenotype. These data expand the knowledge of TSC2 function and pathophysiology which will be highly relevant to future treatments for patients with TSC.

TSC2 inactivation, low mutation burden and high macrophage infiltration characterise hepatic angiomyolipomas.

AIMS: Although TSC1 or TSC2 inactivating mutations that lead to mTORC1 hyperactivation have been reported in hepatic angiomyolipomas (hAML), the role of other somatic genetic events that may contribute to hAML development is unknown. There are also limited data regarding the tumour microenvironment (TME) of hAML. The aim of the present study was to identify other somatic events in genomic level and changes in TME that contribute to tumorigenesis in hAML. METHODS AND RESULTS: In this study, we performed exome sequencing in nine sporadic hAML tumours and deep-coverage targeted sequencing for TSC2 in three additional hAML. Immunohistochemistry and multiplex immunofluorescence were carried out for 15 proteins to characterise the tumour microenvironment and assess immune cell infiltration. Inactivating somatic variants in TSC2 were identified in 10 of 12 (83%) cases, with a median allele frequency of 13.6%. Five to 18 somatic variants (median number: nine, median allele frequency 21%) not in TSC1 or TSC2 were also identified, mostly of uncertain clinical significance. Copy number changes were rare, but detection was impaired by low tumour purity. Immunohistochemistry demonstrated numerous CD68+ macrophages of distinct appearance from Küpffer cells. Multiplex immunofluorescence revealed low numbers of exhausted PD-1+/PD-L1+, FOXP3+ and CD8+ T cells. CONCLUSION: hAML tumours have consistent inactivating mutations in TSC2 and have a low somatic mutation rate, similar to other TSC-associated tumours. Careful histological review, standard IHC and multiplex immunofluorescence demonstrated marked infiltration by non-neoplastic inflammatory cells, mostly macrophages.

Sclerotic Bone Lesions as a Clue in the Diagnosis of Three Generations of Tuberous Sclerosis Complex: Case Report and Review of Literature.

Tuberous sclerosis complex (TSC) is an autosomal dominant genetic disorder that can involve multiple organ systems. Diagnosis is based on independent clinical diagnostic criteria and genetic diagnostic criteria (pathogenic variants on TSC1 and TSC2 genes). To make a definitive diagnosis can be especially difficult in oligosymptomatic or asymptomatic patients and in those patients with genetic variants of uncertain significance (VUS). Early diagnosis and lifelong surveillance are paramount to avoid morbidity and potentially life-threatening complications. To increase diagnostic sensibility, less known manifestations of TSC can be helpful. Herein we show a case in which SBLs were used as a diagnostic clue to help diagnose three generations of oligosymptomatic TSC carrying a VUS in TSC1. SBLs are commonly detected in imaging studies of patients with TSC and have been recently included as a minor clinical diagnostic criterion. Clinicians and radiologists should be aware of their significance as they can be mistaken with osteoblastic metastases.

Design of negative-regulating proteins of Rheb/mTORC1 with much-reduced sizes of the tuberous sclerosis protein complex.

The mTORC1 signaling pathway regulates cell growth and metabolism in a variety of organisms from yeast to human, and inhibition of the mTORC1 pathway has the prospect to treat cancer or achieve longevity. The tuberous sclerosis protein complex (TSCC) is a master negative regulator of the mTORC1 signaling pathway through hydrolyzing the GTP loaded on the small GTPase Rheb, which is a key activator of mTOR. However, the large size (~700 kDa) and complex structural organization of TSCC render it vulnerable to degradation and inactivation, thus limiting its potential application. In this work, based on thorough analysis and understanding of the structural mechanism of how the stabilization domain of TSC2 secures the association of TSC2-GAP with Rheb and thus enhances its GAP activity, we designed two proteins, namely SSG-MTM (short stabilization domain and GAP domain-membrane targeting motif) and SSG-TSC1N, which were able to function like TSCC to negatively regulate Rheb and mTORC1, but with much-reduced sizes (~1/15 and ~ 1/9 of the size of TSCC, respectively). Biochemical and cell biological assays demonstrated that these designed proteins indeed could promote the GTPase activity of Rheb to hydrolyze GTP, inhibit the kinase activity of mTORC1, and prevent mTORC1 from down-regulating catabolism and autophagy.

Glutaredoxin-1 promotes lymphangioleiomyomatosis progression through inhibiting Bim-mediated apoptosis via COX2/PGE2/ERK pathway.

BACKGROUND: Lymphangioleiomyomatosis (LAM) is a female-predominant interstitial lung disease, characterized by progressive cyst formation and respiratory failure. Clinical treatment with the mTORC1 inhibitor rapamycin could relieve partially the respiratory symptoms, but not curative. It is urgent to illustrate the fundamental mechanisms of TSC2 deficiency to the development of LAM, especially mTORC1-independent mechanisms. Glutaredoxin-1 (Glrx), an essential glutathione (GSH)-dependent thiol-oxidoreductase, maintains redox homeostasis and participates in various processes via controlling protein GSH adducts. Redox signalling through protein GSH adducts in LAM remains largely elusive. Here, we demonstrate the underlying mechanism of Glrx in the pathogenesis of LAM. METHODS: 1. Abnormal Glrx expression in various kinds of human malignancies was identified by the GEPIA tumour database, and the expression of Glrx in LAM-derived cells was detected by real-time quantitative reverse transcription (RT-qPCR) and immunoblot. 2. Stable Glrx knockdown cell line was established to evaluate cellular impact. 3. Cell viability was determined by CCK8 assay. 4. Apoptotic cell number and intracellular reactive oxygen species (ROS) level were quantified by flow cytometry. 5. Cox2 expression and PGE2 production were detected to clarify the mechanism of Bim expression modulated by Glrx. 6. S-glutathionylated p65 was enriched and detected by immunoprecipitation and the direct regulation of Glrx on p65 was determined. 7. The xenograft animal model was established and photon flux was analyzed using IVIS Spectrum. RESULTS: In LAM, TSC2 negatively regulated abnormal Glrx expression and activation in a mTORC1-independent manner. Knockdown of Glrx increased the expression of Bim and the accumulation of ROS, together with elevated S-glutathionylated proteins, contributing to the induction of apoptotic cell death and inhibited cell proliferation. Knockdown of Glrx in TSC2-deficient LAM cells increased GSH adducts on nuclear factor-kappa B p65, which contributed to a decrease in the expression of Cox2 and the biosynthesis of PGE2. Inhibition of PGE2 metabolism attenuated phosphorylation of ERK, which led to the accumulation of Bim, due to the imbalance of its phosphorylation and proteasome degradation. In xenograft tumour models, knockdown of Glrx in TSC2-deficient LAM cells inhibited tumour growth and increased tumour cell apoptosis. CONCLUSIONS: Collectively, we provide a novel redox-dependent mechanism in the pathogenesis of LAM and propose that Glrx may be a beneficial strategy for the treatment of LAM or other TSC-related diseases.

An Integral Approach to the Molecular Diagnosis of Tuberous Sclerosis Complex: The Role of Mosaicism and Splicing Variants.

Tuberous sclerosis complex (TSC) is an autosomal dominant genetic disorder characterized by the presence of hamartomas in multiple organs. At the molecular level, the disease is caused by pathogenic variants in the TSC1 and TSC2 genes, and only 10% to 25% of clinically diagnosed patients remain negative after multiplex ligation-dependent probe amplification and exon sequencing of both genes. Here, to improve the molecular diagnosis of TSC, we developed an integral approach that includes multiplex ligation-dependent probe amplification and deep-coverage next-generation sequencing of the entire TSC1 and TSC2 genes, along with an adapted bioinformatic pipeline to detect variants at low allele frequencies (>1%). Using this workflow, the molecular cause was identified in 29 of 42 patients with TSC, describing here, for the first time, 12 novel pathogenic variants in TSC genes. These variants included seven splicing variants, five of which were studied at the cDNA level, determining their effect on splicing. In addition, 8 of the 29 pathogenic variants were detected in mosaicism, including four patients with previous negative study results who presented extremely low mosaic variants (allele frequency, <16%). We demonstrate that this integral approach allows the molecular diagnosis of patients with TSC and improves the conventional one by adapting the technology to the detection of low-frequency mosaics.

Mild TSC phenotype and non-penetrance associated with a frameshift variant in TSC2 prompts caution in evaluating pathogenicity of frameshift variants.

INTRODUCTION: Technological advances in genetic testing, particularly the adoption of noninvasive prenatal screening (NIPS) for single gene disorders such as tuberous sclerosis complex (TSC, OMIM# 613254), mean that putative/possible pathogenetic DNA variants can be identified prior to the appearance of a disease phenotype. Without a phenotype, accurate prediction of variant pathogenicity is crucial. Here, we report a TSC2 frameshift variant, NM_000548.5(TSC2):c.4255_4256delCA, predicted to result in nonsense-mediated mRNA decay (NMD) and cessation of TSC2 protein production and thus pathogenic according to ACMG criteria, identified by NIPS and subsequently detected in family members with few or no symptoms of TSC. Due to the lack of TSC-associated features in the family, we hypothesized that the deletion created a non-canonical 5' donor site resulting in cryptic splicing and a transcript encoding active TSC2 protein. Verifying the predicted effect of the variant was key to designating pathogenicity in this case and should be considered for other frameshift variants in other genetic disorders. METHODS: Phenotypic information on the family members was collected via review of the medical records and patient reports. RNA studies were performed using proband mRNA isolated from blood lymphocytes for RT-PCR and Sanger sequencing. Functional studies were performed by transient expression of the TSC2 variant proteins in cultured cells, followed by immunoblotting. RESULTS: No family members harboring the variant met any major clinical diagnostic criteria for TSC, though a few minor features non-specific to TSC were present. RNA studies supported the hypothesis that the variant caused cryptic splicing, resulting in an mRNA transcript with an in-frame deletion of 93 base pairs r.[4255_4256del, 4251_4343del], p.[(Gln1419Valfs*104), (Gln1419_Ser1449del)]. Expression studies demonstrated that the canonical function of the resulting truncated TSC2 p.Gln1419_Ser1449del protein product was maintained and similar to wildtype. CONCLUSION: Although most frameshift variants are likely to result in NMD, the NM_000548.5(TSC2):c.4255_4256delCA variant creates a cryptic 5' splice donor site, resulting in an in-frame deletion that retains TSC2 function, explaining why carriers of the variant do not have typical features of TSC. The information is important for this family and others with the same variant. Equally important is the lesson that predictions can be inaccurate, and that caution should be used when designating frameshift variants as pathogenic, especially when phenotypic information to corroborate testing results is unavailable. Our work demonstrates that functional RNA- and protein-based confirmation of the effects of DNA variants improves molecular genetic diagnostics.

Establishment of human induced pluripotent stem cell lines, KMUGMCi006, from a patient with Tuberous sclerosis complex (TSC) bearing mosaic nonsense mutations in the Tuberous sclerosis complex 2 (TSC2) gene.

Tuberous sclerosis complex (TSC) is an autosomal dominant disorder characterized by neuropsychiatric symptoms and multiple dysplastic organ lesions, caused by loss of function mutations in either TSC1 or TSC2. The peripheral blood mononuclear cells (PBMCs) from a patient carrying mosaic nonsense mutation of TSC2 gene were reprogrammed using the CytoTune-iPS2.0 Sendai Reprogramming Kit. The human induced pluripotent cell (hiPSC) lines with the mutation and without the mutation were established. The heterozygous nonsense mutation in TSC2 will cause the truncated protein, which is known to associated with TSC. The established hiPSC lines will enable proper in vitro disease modelling of TSC.

Comprehensive genetic and phenotype analysis of 95 individuals with mosaic tuberous sclerosis complex.

Tuberous sclerosis complex (TSC) is a neurogenetic disorder due to loss-of-function TSC1 or TSC2 variants, characterized by tumors affecting multiple organs, including skin, brain, heart, lung, and kidney. Mosaicism for TSC1 or TSC2 variants occurs in 10%-15% of individuals diagnosed with TSC. Here, we report comprehensive characterization of TSC mosaicism by using massively parallel sequencing (MPS) of 330 TSC samples from a variety of tissues and fluids from a cohort of 95 individuals with mosaic TSC. TSC1 variants in individuals with mosaic TSC are much less common (9%) than in germline TSC overall (26%) (p < 0.0001). The mosaic variant allele frequency (VAF) is significantly higher in TSC1 than in TSC2, in both blood and saliva (median VAF: TSC1, 4.91%; TSC2, 1.93%; p = 0.036) and facial angiofibromas (median VAF: TSC1, 7.7%; TSC2 3.7%; p = 0.004), while the number of TSC clinical features in individuals with TSC1 and TSC2 mosaicism was similar. The distribution of mosaic variants across TSC1 and TSC2 is similar to that for pathogenic germline variants in general TSC. The systemic mosaic variant was not present in blood in 14 of 76 (18%) individuals with TSC, highlighting the value of analysis of multiple samples from each individual. A detailed comparison revealed that nearly all TSC clinical features are less common in individuals with mosaic versus germline TSC. A large number of previously unreported TSC1 and TSC2 variants, including intronic and large rearrangements (n = 11), were also identified.

Estradiol Augments Tumor-Induced Neutrophil Production to Promote Tumor Cell Actions in Lymphangioleiomyomatosis Models.

Lymphangioleiomyomatosis (LAM) is a rare cystic lung disease caused by smooth muscle cell-like tumors containing tuberous sclerosis (TSC) gene mutations and found almost exclusively in females. Patient studies suggest LAM progression is estrogen dependent, an observation supported by in vivo mouse models. However, in vitro data using TSC-null cell lines demonstrate modest estradiol (E2) responses, suggesting E2 effects in vivo may involve pathways independent of direct tumor stimulation. We previously reported tumor-dependent neutrophil expansion and promotion of TSC2-null tumor growth in an E2-sensitive LAM mouse model. We therefore hypothesized that E2 stimulates tumor growth in part by promoting neutrophil production. Here we report that E2-enhanced lung colonization of TSC2-null cells is indeed dependent on neutrophils. We demonstrate that E2 induces granulopoiesis via estrogen receptor α in male and female bone marrow cultures. With our novel TSC2-null mouse myometrial cell line, we show that factors released from these cells drive E2-sensitive neutrophil production. Last, we analyzed single-cell RNA sequencing data from LAM patients and demonstrate the presence of tumor-activated neutrophils. Our data suggest a powerful positive feedback loop whereby E2 and tumor factors induce neutrophil expansion, which in turn intensifies tumor growth and production of neutrophil-stimulating factors, resulting in continued TSC2-null tumor growth.

Pathogenic RHEB Somatic Variant in a Child With Tuberous Sclerosis Complex Without Pathogenic Variants in TSC1 or TSC2.

OBJECTIVE: To describe a child meeting diagnostic criteria for tuberous sclerosis complex (TSC) carrying a pathogenic somatic variant in RHEB, but no pathogenic variants in the 2 known TSC genes, TSC1 or TSC2. METHODS: We present the clinical and imaging findings in a child presenting with drug-resistant focal seizures and multiple cortical tubers, a subependymal giant cell astrocytoma and multiple subependymal nodules in 1 cerebral hemisphere. Targeted panel sequencing and exome sequencing were performed on genomic DNA derived from blood and resected tuber tissue. RESULTS: The child satisfied clinical diagnostic criteria for TSC, having 3 major features, only 2 of which are required for diagnosis. Genetic testing did not identify pathogenic variants or copy number variations in TSC1 or TSC2 but identified a pathogenic somatic RHEB variant (NM_005614.4:c.104_105delACinsTA [p.Tyr35Leu]) in the cortical tuber. DISCUSSION: RHEB is a partner of the TSC1/2 complex in the mechanistic target of rapamycin pathway. Somatic variants in RHEB are associated with focal cortical dysplasia and hemimegalencephaly. We propose that variants in RHEB may explain some of the genetically undiagnosed TSC cases and may be the third gene for TSC, or TSC3.

Review: Mammalian Target of Rapamycin (mTOR) Pathway Is Critical in Developing Most Renal Cell Tumors.

OBJECTIVE: Various renal cell carcinomas (RCC) are derived from different segments of the renal tubular origin, which determines their morphological and immunohistochemical phenotype and their molecular signaling pathway as a therapeutic target. Most of these tumors utilize the mammalian target of rapamycin (mTOR) pathway to activate pathways involving metabolic and nutritional supplies. METHODS: Overexpressed mTOR signals are reported in more than 90% of the most common types of RCC. Many new renal tumor entities have been reported in recent years. RESULTS: Among them, somatic mutations in tuberous sclerosis complex (TSC) result in loss of its normal inhibitory control over mTOR, thus promoting mTOR-associated proliferative activities in several new renal neoplastic entities including RCC with fibromyomatous stroma (RCCFMS), eosinophilic vacuolated tumor, eosinophilic solid & cystic RCC, and low-grade oncocytic tumor. CONCLUSIONS: This short review provides a comprehensive correlation of tumor morphology and immunohistochemical phenotype with renal tubular differentiation and their shared mTOR. These essential pieces of knowledge are vital in the diagnosis and clinical management of renal cell neoplasms.

Reprint of: lessons from histopathologic examination of nephrectomy specimens in patients with tuberous sclerosis complex: cysts, angiomyolipomas & renal cell carcinoma.

Renal manifestations in patients with tuberous sclerosis complex (TSC) include cysts, angiomyolipoma, and renal cell carcinoma. Unlike many hereditary predisposition syndromes, the spectrum of renal tumors in TSC patients (including both angiomyolipoma and renal cell carcinoma) is broad, with significant morphologic heterogeneity. An improved understanding of histopathologic findings in TSC patients and associated clinicopathologic correlates has significant implications not just in establishing a diagnosis of TSC, but also in the recognition of sporadic tumors occurring secondary to somatic alterations of TSC1/TSC2/MTOR pathway genes and accurate prognostication. In this review, we have discussed issues relevant to clinical management based on histopathologic findings in nephrectomy specimens from patients with TSC. This includes discussions related to screening for TSC, diagnosis of PKD1/TSC2 contiguous gene deletion syndrome, the morphologic spectrum of angiomyolipoma and renal epithelium-derived neoplasia, including the risk of disease progression.

Refractive Errors, Retinal Findings, and Genotype of Tuberous Sclerosis Complex: A Retrospective Cohort Study.

PURPOSE: To examine the refractive errors, retinal manifestations, and genotype in tuberous sclerosis complex (TSC) patients in a Korean population. MATERIALS AND METHODS: A total of 98 patients with TSC were enrolled in Severance Hospital for a retrospective cohort study. The number of retinal astrocytic hamartoma and retinal achromic patch within a patient, as well as the size, bilaterality, and morphological type were studied. In addition, the refractive status of patients and the comorbidity of intellectual disability and epilepsy were also examined. RESULTS: Retinal astrocytic hamartoma was found in 37 patients, and bilateral invasion was observed in 20 patients (54%). TSC1 mutation was associated with myopia (p=0.01), while TSC2 mutation was associated with emmetropia (p=0.01). Retinal astrocytic hamartoma was categorized into three morphological types and examined as follows: type I (87%), type II (35%), and type III (14%). Single invasion of retinal astrocytic hamartoma was identified in 32% of the patients, and multiple invasions in 68%. The TSC1/TSC2 detection rate was 91% (41/45). Among them, TSC1 variant was detected in 23 patients (54%), whereas TSC2 variant was detected in 18 patients (40%). The results showed that TSC2 mutations are correlated with a higher rate of retinal astrocytic hamartoma involvement (all p<0.05), and multiple and bilateral involvement of retinal hamartomas (all p<0.05). However, the size of retinal astrocytic hamartomas, comorbidity of epilepsy, or intellectual disability did not show correlation with the genetic variant. CONCLUSION: TSC1 variant patients were more myopic, while TSC2 variant patients showed association with more extensive involvement of retinal astrocytic hamartoma.

TSC2 regulates tumor susceptibility to TRAIL-mediated T-cell killing by orchestrating mTOR signaling.

Resistance to cancer immunotherapy continues to impair common clinical benefit. Here, we use whole-genome CRISPR-Cas9 knockout data to uncover an important role for Tuberous Sclerosis Complex 2 (TSC2) in determining tumor susceptibility to cytotoxic T lymphocyte (CTL) killing in human melanoma cells. TSC2-depleted tumor cells had disrupted mTOR regulation following CTL attack, which was associated with enhanced cell death. Wild-type tumor cells adapted to CTL attack by shifting their mTOR signaling balance toward increased mTORC2 activity, circumventing apoptosis, and necroptosis. TSC2 ablation strongly augmented tumor cell sensitivity to CTL attack in vitro and in vivo, suggesting one of its functions is to critically protect tumor cells. Mechanistically, TSC2 inactivation caused elevation of TRAIL receptor expression, cooperating with mTORC1-S6 signaling to induce tumor cell death. Clinically, we found a negative correlation between TSC2 expression and TRAIL signaling in TCGA patient cohorts. Moreover, a lower TSC2 immune response signature was observed in melanomas from patients responding to immune checkpoint blockade. Our study uncovers a pivotal role for TSC2 in the cancer immune response by governing crosstalk between TSC2-mTOR and TRAIL signaling, aiding future therapeutic exploration of this pathway in immuno-oncology.