TSC1 blocking peptide

BACKGROUND: Lymphangioleiomyomatosis (LAM) is a destructive metastasizing neoplasm of the lung characterized by proliferation of LAM cells in specialized lung nodules. LAM cells are characterized by expression of the prometastatic and cancer-initiating hyaluronan receptor CD44v6, and loss of heterozygosity (LOH) of TSC1 and TSC2. The circulating neoplastic LAM cells are thought to be involved in metastasis. Because LAM cells display properties of neoplastic, metastatic, and stem cell-like cancer cells, we hypothesized that elevated aldehyde dehydrogenase (ALDH) activity, characteristic of cancer and stem cells, is a property of LAM cells. METHODS: We performed an in silico search of ALDH genes in microdissected LAM lung nodules. To identify circulating LAM cells, we osmotically removed red blood cells from whole blood to obtain peripheral blood mononuclear cells, which were then sorted by fluorescence-activated cell sorting based on their level of ALDH activity. RESULTS: Microdissected LAM lung nodules possess a distinctive ALDH gene profile. The cell subpopulation with high ALDH activity, isolated from circulating cells, possessed TSC2 LOH in 8 of 14 patients with LAM. Approximately 60% of the circulating cells with high ALDH activity expressed CD44v6. Cells with TSC2 LOH from patients with LAM and LAM/TSC exhibited different properties in different body locations, but all cell types showed high ALDH activity. CONCLUSIONS: This new procedure allows for isolation of circulating LAM cells from cultured cells, blood, and chylous effusions and shows that circulating LAM cells are heterogeneous with neoplastic, metastatic, and cancer-stem cell-like properties.

Rapamycins are immunosuppressant and anti-cancer drugs that inhibit the kinase mTOR. Clinically, they often cause bone pain, bone necrosis, and high bone turnover, yet the mechanisms are unclear. Here we show that mTORC1 activity is high in osteoclast precursors but downregulated upon RANKL treatment. Loss-of-function genetic models reveal that while early Raptor deletion in hematopoietic stem cells blunts osteoclastogenesis due to compromised proliferation/survival, late Raptor deletion in osteoclast precursors instead augments osteoclastogenesis. Gain-of-function genetic models by TSC1 deletion in HSCs or osteoclast precursors cause constitutive mTORC1 activation, impairing osteoclastogenesis. Pharmacologically, rapamycin treatment at low but clinically relevant doses exacerbates osteoclast differentiation and bone resorption, leading to bone loss. Mechanistically, RANKL inactivates mTORC1 via calcineurin-mediated mTORC1 dephosphorylation, consequently activating NFATc1 by reducing mTORC1-mediated NFATc1 phosphorylation. These findings uncover biphasic roles of mTORC1 in osteoclastogenesis, dosage-dependent effects of rapamycin on bone, and a previously unrecognized calcineurin-mTORC1-NFATc1 phosphorylation-regulatory signaling cascade.