STUDIO BIOCHIMICO E FARMACOLOGICO DI CELLULE MUSCOLARI LISCE ISOLATE DA TESSUTI DI PAZIENTI AFFETTI DA TSC2 E LAM. MODIFICAZIONI EPIGENETICHE DEL GENE TSC2 NELLA PATOGENENESI DELLA TSC E DELLA LAM.

Tuberous sclerosis complex (TSC) and lymphangioleiomymatosis (LAM) are rare diseases. TSC is a genetic disease caused by mutation in TSC1 or TSC2 genes. TSC2 cells form hamartomas and might invade lungs causing the fatal diseases LAM. From an angiomylipoma (AML) of a TSC male patient we isolated a TSC2 smooth muscle (ASM) population. The growth of these cells was EGF-dependent. They showed a constitutive S6 phosphorylation, and lacked tuberin as the result of TSC2 promoter methylation (TSC2-/meth ASM cells). Chromatin remodeling agents, such as trichostatin–A and 5-azacytidine de-methylated TSC2-/meth ASM cell promoter and induced tuberin expression. The blockade of EGF-receptor with specific antibodies results in cell death as shown in TSC2-/- ASM cells, a population previously isolated from an AML of female TSC patient. LAM cells migrate or metastasize to other organs, in fact cells with TSC2 mutation have been found in AMLs and lung lesions of LAM patients. We isolated a population from chylous of a TSC/LAM patient, in which, as in TSC2-/meth cells, tuberin expression was induced by 5-azacytidine and trichostatin A treatments. These cells required the supplementation of EGF for proliferation, such as TSC2-/- and TSC2-/meth ASM cells. Chylous TSC/LAM cells expressed marker for identification of mesenchimal characteristics, such as vimentin and SNAIL, while E-cadherin, usually not expressed in metastatized cancer cells, was not detectable. The acquisition of mesenchymal characteristics for cancer cells is a transient event that might be important for migration and tissue invasion. Chylous TSC/LAM cells switched from a short floating stage, with low S6 phosphorylation levels, to a stage of adhesion with high S6 phosphorylation. These data suggest a novel therapeutic approach for the control of TSC and LAM abnormal cell growth using anti-EGFR antibody in addition to rapamycin, and, in some cases, the chromatin remodelling agents. Moreover, our data confirm the evidence that TSC2 pathogenesis might originate from an epigenetic defect.