mTOR is a highly conserved serine/threonine kinase that regulates cell growth and meta bolism in response to environmental factors. It is activated downstream of the PI3-K/AKT pathway and executes its biologic functions as two distinct complexes, mTORC1 and mTORC2. mTOR, mLST8/GβL and the negative regulator Deptor are shared by both complexes [1
]. mTORC1 is uniquely composed by the regulatory-associated protein of mTOR (raptor) and the proline-rich AKT substrate of 40 kDa (PRAS40), and is inhibited by rapamycin and the other rapalogs (everolimus and temsirolimus). mTORC2 includes rapamycin insensitive companion of TOR (rictor) and, in contrast to mTORC1, is largely resistant to inhibition by the rapalogs.
The mTOR pathway is most typically activated downstream of the PI3-K/Akt pathway in response to growth factors signaling through tyrosine kinase receptors (e.g., the binding of IGF to its receptor) (). PI3-K mediates the generation of PIP3 from PIP2, an action which is opposed by the tumor suppressor PTEN. PIP3 binds directly to the PH domain of Akt and mediates its localization to the cell membrane where it is then activated by phosphorylation at two sites, threonine 308 and serine 473. One of the many functions of activated Akt is to directly phosphorylate tuberous sclerosis complex 2 (TSC2; tuberin) resulting in the inhibition of its GAP activity towards the small GTPase Rheb. Relief of this GAP activity enhances the GTP loading of Rheb, promoting its activity. Rheb has been shown to bind directly to the kinase domain of mTOR and activate mTOR kinase activity in a GTP-dependent manner [2
mTORC1 acts through its downstream effectors, the S6K and the eukaryotic elongation factor 4E-BP, to regulate cell growth and proliferation in response to growth factors (i.e., IGF), nutrients (amino acids in particular), energy level and environmental stress (e.g., hypoxia, DNA damage and reducing conditions) [2
]. The activation of S6K by mTOR is critical for ribosomal biogenesis [3
], cell growth, anti-apoptosis and translation of the structured 5′ untranslated region (UTR) containing mRNA species, while the phosphorylation (and inactivation) of 4E-BP1 promotes cap-dependent translation. Phosphorylated 4E-BP1 releases eIF4E, a protein that binds the 5′ m7
G cap structure of cellular mRNAs and facilitates translation by enhancing the association of the mRNA with the RNA helicase eIF4A and the ribosome-interacting scaffolding protein eIF4G [4
]. The extent to which the translation of a particular transcript depends on mTORC1 (and eIF4E) activity is determined by the length and complexity of the mRNA 5′UTR. Approximately 3% of cellular mRNAs have 5′UTRs that extend to greater than a third of the transcript length. The 5′UTRs of these mRNAs often have high GC content and stem loop structures that preclude translation, except when mTORC1 is activated. Many of these ‘weak’, otherwise untranslated mRNAs, encode proteins essential for cell cycle progression (e.g., cyclins, c-Myc and ornithine decarboxylase), survival (e.g., the IAPs XIAP and survivin), and angiogenesis (e.g., VEGF and FGF-2) [5
]. Not surprisingly, Dowling et al
. have recently shown that phosphorylation of 4E-BP1 (and resultant activation of eIF4E) is the critical determinant in driving cell proliferation in response to TORC1 activation [6
]. Furthermore, overexpression of eIF4E has been shown to be sufficient to drive malignant transformation in some cells [7
]. Thus, while the precise mechanism of the therapeutic efficacy of mTOR inhibitors remains unknown, it is possible that attenuation of the translation of critical gene products driven by 4E-BP1 phosphorylation (and resultant eIF4E activation) by mTORC1 may be an important aspect of this effect.
The mTOR pathway may be of particular relevance to RCC as it has been shown that HIF protein expression is dependent on mTOR in certain cellular contexts. Inappropriate accumulation of HIF-1α and HIF-2α as a result of biallelic alterations in the von Hippel-Lindau (VHL
) gene observed in the majority of clear cell RCC is believed to be a critical step in RCC tumorigenesis as a result of increased expression of HIF-regulated gene products including VEGF, PDGF and TGF-α. Toschi et al
. have shown that mTOR activation, in conjunction with the presence of phospholipase D, enhances the expression of both HIF-1α and HIF-2α in RCC cells at a translational level rather than transcriptional [8
]. Furthermore, treatment of mice bearing RCC xenograft models with temsirolimus has been shown to result in impaired expression of HIF-1α under both hypoxic and normoxic conditions, presenting another potential mechanism of action for the rapalogs in patient with RCC [9