u-0126 and Tuberous-Sclerosis

Tuberous sclerosis complex (TSC) is an autosomal dominantly inherited neurocutaneous disorder caused by inactivating mutations in TSC1 or TSC2, key regulators of the mechanistic target of rapamycin complex 1 (mTORC1) pathway. In the CNS, TSC is characterized by cortical tubers, subependymal nodules and subependymal giant cell astrocytomas (SEGAs). SEGAs may lead to impaired circulation of CSF resulting in hydrocephalus and raised intracranial pressure in patients with TSC. Currently, surgical resection and mTORC1 inhibitors are the recommended treatment options for patients with SEGA. In the present study, high-throughput RNA-sequencing (SEGAs n = 19, periventricular control n = 8) was used in combination with computational approaches to unravel the complexity of SEGA development. We identified 9400 mRNAs and 94 microRNAs differentially expressed in SEGAs compared to control tissue. The SEGA transcriptome profile was enriched for the mitogen-activated protein kinase (MAPK) pathway, a major regulator of cell proliferation and survival. Analysis at the protein level confirmed that extracellular signal-regulated kinase (ERK) is activated in SEGAs. Subsequently, the inhibition of ERK independently of mTORC1 blockade decreased efficiently the proliferation of primary patient-derived SEGA cultures. Furthermore, we found that LAMTOR1, LAMTOR2, LAMTOR3, LAMTOR4 and LAMTOR5 were overexpressed at both gene and protein levels in SEGA compared to control tissue. Taken together LAMTOR1-5 can form a complex, known as the 'Ragulator' complex, which is known to activate both mTORC1 and MAPK/ERK pathways. Overall, this study shows that the MAPK/ERK pathway could be used as a target for treatment independent of, or in combination with mTORC1 inhibitors for TSC patients. Moreover, our study provides initial evidence of a possible link between the constitutive activated mTORC1 pathway and a secondary driver pathway of tumour growth.

Topics: Adaptor Proteins, Signal Transducing; Adolescent; Adult; Astrocytes; Astrocytoma; Brain Neoplasms; Butadienes; Child; Child, Preschool; Enzyme Inhibitors; Extracellular Signal-Regulated MAP Kinases; Female; Gene Expression Profiling; Guanine Nucleotide Exchange Factors; High-Throughput Nucleotide Sequencing; Humans; Infant; Intracellular Signaling Peptides and Proteins; Male; MAP Kinase Signaling System; Mechanistic Target of Rapamycin Complex 1; MicroRNAs; Nitriles; RNA-Seq; RNA, Messenger; Sequence Analysis, RNA; Tuberous Sclerosis; Tuberous Sclerosis Complex 1 Protein; Tuberous Sclerosis Complex 2 Protein; Tumor Cells, Cultured; Young Adult

Tuberous sclerosis (TS), neurological disorder manifesting with the formation of tumors in numerous organ systems, is a disease associated with the upregulation of mammalian target of rapamycin (mTOR) pathway. It has been found that in healthy individuals two tumor suppressor genes, TSC1 and TSC2, encoding proteins called hamartin and tuberin, respectively, are responsible for the control over mTOR kinase. Loss of one of these genes constitutes the genetic background of TS. In the current study, we aimed at evaluating the fitness of the only TS-associated sarcoma cell line deposited in American Tissue Culture Collection, TSC2ang1, for the in vitro studies on TS. We found that the line shows a stable chromosome pattern with typical Robertsonian translocations. Similarly to primary tumors from TS patients, TSC2ang1 cells respond to rapamycin-induced mTOR inhibition. The cells demonstrate activation of both Akt and Erk pathways, but inhibition of neither of them is as effective as mTOR suppression when considering proliferation potential. Based on these results we propose TSC2ang1 as a good and stable model for pathophysiological and pharmacological studies on skin lesions in TS.

Topics: Androstadienes; Animals; Butadienes; Cell Line; Cell Proliferation; CHO Cells; Chromones; Chromosomes, Human; Cricetinae; Cricetulus; Enzyme Activation; Extracellular Signal-Regulated MAP Kinases; Heterozygote; Humans; Karyotyping; Morpholines; Nitriles; Phosphoinositide-3 Kinase Inhibitors; Protein Kinases; Proto-Oncogene Proteins c-akt; Skin Neoplasms; Substrate Specificity; TOR Serine-Threonine Kinases; Tuberous Sclerosis; Tuberous Sclerosis Complex 2 Protein; Tumor Suppressor Proteins; Wortmannin

Tuberous sclerosis (TS) is an autosomal dominant disease associated with the formation of usually benign tumors or hamartomas. The disease is connected with upregulation of mammalian target of rapamycin, central regulator of protein translation, which is usually regarded to be activated by Akt kinase. Here, we show for the first time that in all four brain lesions and one angiomyolipoma from TS patients both extracellular signal-regulated kinase (Erk) and p90 ribosomal S6 kinase 1 activation as well as Erk-dependent phosphorylation of p70 ribosomal S6 kinase 1 are markedly elevated whereas Akt, participating in the classical pathway of mammalian target of rapamycin activation is not always activated. Erk activation is also present in TS-derived cell lines. Importantly, Erk inhibition leads to the decrease of proliferation potential of such lines. These results show that Erk is specifically implicated in the pathogenesis of hamartomas.

Topics: Angiomyolipoma; Animals; Astrocytoma; Brain Neoplasms; Butadienes; Cell Line; Enzyme Activation; Enzyme Inhibitors; Extracellular Signal-Regulated MAP Kinases; Humans; Nitriles; Protein Kinases; Proto-Oncogene Proteins c-akt; TOR Serine-Threonine Kinases; Tuberous Sclerosis