and also in human cancer. in fact, they have been proven to be more potent than rapalogs. theoretically, the most important advantages of these mtor inhibitors is the considerable decrease of akt phosphorylation on mtorc2 blockade and in addition to a better inhibition on mtorc1. however, some drawbacks exist. even though these compounds have been effective in rapamycin-insensitive cell lines, they have only shown limited success in kras driven tumors. this suggests that combinational therapy may be necessary for the treatment of these cancers. another drawback is also their potential toxicity. these facts have raised concerns about the long term efficacy of these types of inhibitors. the close interaction of mtor with the pi3k pathway has also led to the development of mtor/pi3k dual inhibitors. compared with drugs that inhibit either mtorc1 or pi3k, these drugs have the benefit of inhibiting mtorc1, mtorc2, and all the catalytic isoforms of pi3k. targeting both kinases at the same time reduces the upregulation of pi3k, which is typically produced with an inhibition on mtorc1. the inhibition of the pi3k/mtor pathway has been shown to potently block proliferation by inducing g1 arrest in different tumor cell lines. strong induction of apoptosis and autophagy has also been seen. despite good promising results, there are preclinical evidence that some types of cancers may be insensitive to this dual inhibition. the dual pi3k/mtor inhibitors are also likely to have increased toxicity. mechanism of action the studies of rapamycin as immunosuppressive agent enabled us to understand its mechanism of action. it inhibits t-cell proliferation and proliferative responses induced by several cytokines, including interleukin 1 (il-1), il-2, il-3, il-4, il-6, igf, pdgf, and colony-stimulating factors (csfs). rapamycin inhibitors and rapalogs can target tumor growth both directly and indirectly. direct impact of them on cancer cells depend on the concentration of the drug and certain cellular characteristics. the indirect way, is based on interaction with processes required for tumor angiogenesis. effects in cancer cells effects of rapamycin and rapalogs in cancer cells rapamycin and rapalogs crosslink the immunophilin fk506 binding protein, tacrolimus or fkbp-12, through its methoxy group. the rapamycin-fkbp12 complex interferes with frb domain of mtor. molecular interaction between fkbp12, mtor, and rapamycin can last for about three days (72 hours). the inhibition of mtor blocks the binding of the accessory protein raptor (regulatory-associated protein of mtor) to mtor, but that is necessary for downstream phosphorylation of s6k1 and 4ebp1. as a consequence, s6k1 dephosphorylates, which reduces protein synthesis and decreases cell mortality and size. rapamycin induces dephosphorylation of 4ebp1 as well, resulting in an increase in p27 and a decrease in cyclin d1 expression. that leads to late blockage of g1/s cell cycle. rapamycin has shown to induce cancer cell death by stimulating autophagy or apoptosis, but the molecular mechanism of apoptosis in cancer cells has not yet been fully resolved. one suggestion of the relation between mtor inhibition and apoptosis might be through the downstream target s6k1, which can phosphorylate bad, a pro-apoptotic molecule, on ser136. that reaction breaks the binding of bad to bcl-xl and bcl2, a mitochondrial death inhibitors, resulting in inactivation of bad and decreased cell survival. rapamycin has also shown to induce p53-independent apoptosis in certain types of cancer. effects on tumor angiogenesis effects of rapamycin and rapalogs on endothelial and tumor cells tumor angiogenesis rely on interactions between endothelial vascular growth factors which can all activate the pi3k/akt/mtor in endothelial cells, pericytes, or cancer cells. example of these growth factors are angiopoietin 1 (ang1), ang 2, basic fibroblast growth factor (bfgf), ephrin-b2, vascular endothelial growth factor (vegf), and members of the tumor growth factor-β (tgfβ) superfamily. one of the major stimuli of angiogenesis is hypoxia, resulting in activation of hypoxia-inducible transcription factors (hifs) and expression of ang2, bfgf, pdgf, vegf, and vegfr. inhibition of hif1α translation by preventing pdgf/pdgfr and vegf/vegfr can result from mtor inhibition. a g0-g1 cell-cycle blockage can be the consequence of inactivation of mtor in hypoxia-activated pericytes and endothelial cells. there is some evidence that extended therapy with rapamycin may have effect on akt and mtorc2 as well. effects on chemotherapy pharmacologic down-regulation of (mtor) pathway during chemotherapy in a mouse model prevents activation of primordial follicles, preserves ovarian function, and maintains normal fertility using clinically available inhibitors ink and rad. in that way, it helps to maintain fertility while undergoing chemotherapy treatments. these mtor inhibitors, when administrated as pretreatment or co-treatment with standard gonadotoxic chemotherapy, helps to maintain ovarian follicles in their primordial state. effects on cognition mtor promotes the protein synthesis required for synaptic plasticity. studies in cell cultures and hippocampal slices indicate that mtor inhibition reduces long-term potentiation. mtor activation can protect against certain neurodegeneration associated with certain disease conditions. on the other hand, promotion of autophagy by mtor inhibition may reduce cognitive decline associated with neurodegeneration. moderate reduction of mtor activity by 25-30% has been shown to improve brain function, suggesting that the relation between mtor and cognition is optimized with intermediate doses (2.24 mg/kg/day in mice, human equivalent about 0.19 mg/kg/day), where very high or very low doses impair cognition. reduction of the inflammatory cytokine interleukin 1 beta (il-1β) in mice by mtor inhibition (with rapamycin in doses of 20 mg/kg/day, human equivalent about 1.6 mg/kg/day) has been shown to enhance learning and memory. although il-1β is required for memory, il-1β normally increases with age, impairing cognitive function. structure activity relationship the pipecolate region of rapamycin structure seems necessary for rapamycin-binding to fkbp12. this step is required for further binding of rapamycin to the mtor kinase, which is the key enzyme in many biological actions of rapamycin. the high affinity of rapamycin binding to fkbp12 is explained by number of hydrogen bonds through two different hydrophobic binding pockets, and this has been revealed by x-ray crystal structure of the compound bound to the protein. the structural characteristics common to temsirolimus and sirolimus; the pipecolic acid, tricarbonyl region from c13-c15, and lactone functionalities play the key role in binding groups with the fkbp12. the most important hydrogen bonds are the lactone carbonyl oxygen at c-21 to the backbone nh of ile56, amide carbonyl at c-15 to the phenolic group on the sidechain of tyr82, and the hydroxyl proton at the hemiketal carbon, c-13, to the sidechain of asp37. binding sites of rapamycin: green ring (pipecolate region) represents binding region to fkbp12 and purple ring represents binding region to mtor structural changes to the rapamycin structure can affect binding to mtor. this could include both direct and indirect binding as a part of binding to fkbp12. interaction of the fkbp12-rapamycin complex with mtor corresponds with conformational flexibility of the effector domain of rapamycin. this domain consists of molecular regions that make hydrophobic interactions with the fkb domain and triene region from c-1-c-6, methoxy group at c-7, and methyl groups at c-33, c-27 and c-25. all changes of the macrolide ring can have unpredictable effects on binding and therefore, make determination of sar for rapalogs problematic. rapamycin contains no functional groups that ionize in the ph range 1-10 and therefore, are rather insoluble in water. despite its effectiveness in preclinic cancer models, its poor solubility in water, stability, and the long half-life elimination made its parenteral use difficult, but the development of soluble rapamycin analogs vanquished various barriers. nonetheless, the rapamycin analogs that have been approved for human use are modified at c-43 hydroxyl group and show improvement in pharmacokinetic parameters as well as drug properties, for example, solubility. rapamycin and temsirolimus have similar chemical structures and bind to fkbp12, though their mechanism of action differs. temsirolimus is a dihydroxymethyl propionic acid ester of rapamycin, and its first derivative. therefore, it is more water-soluble, and due to its water solubility it can be given by intravenous formulation. everolimus has o-2 hydroxyethyl chain substitution and deforolimus has a phosphine oxide substitution at position c-43 in the lactone ring of rapamycin. deforolimus (ridaforolimus ) has c43 secondary alcohol moiety of the cyclohexyl group of rapamycin that was substituted with phosphonate and phosphinate groups, preventing the high-affinity binding to mtor and fkbp. computational modelling studies helped the synthesise of the compound. adverse events treatment with mtor inhibitors can be complicated by adverse events. the most frequently occurring adverse events are stomatitis, rash, anemia, fatigue, hyperglycemia/hypertriglyceridemia, decreased appetite, nausea, and diarrhea. additionally, interstitial lung disease is an adverse event of particular importance. mtori-induced ild often is asymptomatic (with ground glass abnormalities on chest ct) or mild symptomatic (with a non-productive cough), but can be very severe as well. even fatalities have been described. careful diagnosis and treatment, therefore, is essential. recently, a new diagnostic and therapeutic management approach has been proposed. biomarkers identification of predictive biomarkers of efficacy for tumor types that are sensitive to mtor inhibitors remains a major issue. possible predictive biomarkers for tumor response to mtor inhibitors, as have been described in glioblastoma, breast and prostate cancer cells, may be the differential expression of mtor pathway proteins, pten, akt, and s6. thus, this data is based on preclinical assays, based on in vitro cultured tumor cell lines, which suggest that the effects of mtor inhibitors may be more pronounced in cancers displaying loss of pten functions or pik3ca mutations. however, the use of pten, pik3ca mutations, and akt–phospho status for predicting rapalog sensitivity has not been fully validated in clinic. to date, attempts to identify biomarkers of rapalog response have been unsuccessful. sensitivity clinical and translational data suggest that sensitive tumor types, with adequate parameters and functional apoptosis pathways, might not need high doses of mtor inhibitors to trigger apoptosis. in most cases, cancer cells might only be partially sensitive to mtor inhibitors due to redundant signal transduction or lack of functional apoptosis signaling pathways. in situations like this, high doses of mtor inhibitors might be required. in a recent study of patients with renal cell carcinoma, resistance to temsirolimus was associated with low levels of p-akt and p-s6k1, that play the key role in mtor activation. these data strongly suggests number of tumors with an activated pi3k/akt/mtor signaling pathway that does not respond to mtor inhibitors. for future studies, it is recommended to exclude patients with low or negative p-akt levels from trials with mtor inhibitors. current data is insufficient to predict sensitivity of tumors to rapamycin. however, the existing data allows us to characterize tumors that might not respond to rapalogs. atp-competitive mtor kinase inhibitors these second generation mtor inhibitors bind to atp-binding site in mtor kinase domain required for the functions of both mtorc1 and mtorc2, and result in downregulation of mtor signaling pathway. due to pi3k and mtorc2 ability to regulate akt phosphorylation, these two compounds play a key role in minimizing the feedback activation of akt. mtor/pi3k dual inhibitors several, so-called mtor/pi3k dual inhibitors (tpdis), have been developed and are in early-stage preclinical trials and show promising results. their development has been benefited from previous studies with pi3k-selective inhibitors. the activity of these small molecules from rapalog activity differs in the way by blocking both mtorc1-dependent phospholylation of s6k1 and mtorc2-dependent phosphorylation of akt ser473 residue. dactolisib chemical structure dual mtor/pi3k inhibitors include dactolisib, voxtalisib, bgt226, sf1126, pki-587 and many more. for example, novartis has developed the compound nvpbe235 that was reported to inhibit tumor growth in various preclinical models. it enhances antitumor activity of some other drugs such as vincristine. dactolisib seems to inhibit effectively both wild-type and mutant form of pi3kca, which suggests its use towards wide types of tumors. studies have shown superior antiproliferative activity to rapalogs and in vivo models have confirmed these potent antineoplastic effects of dual mtor/pi3k inhibitors. these inhibitors target isoforms of pi3k (p110α, β and γ) along with atp-binding sites of mtorc1 and mtorc2 by blocking pi3k/akt signaling, even in cancer types with mutations in this pathway. mtorc1/mtorc2 dual inhibitors (torcdis) sapanisertib (ink-128): chemical structure new mtor-specific inhibitors came forth from screening and drug discovery efforts. these compounds block activity of both mtor complexes and are called mtorc1/mtorc2 dual inhibitors. compounds with this characteristics such as sapanisertib (codenamed ink128), azd8055, and azd2014 have entered clinical trials. a series of these mtor kinase inhibitors have been studied. their structure is derived from morpholino pyrazolopyrimidine scaffold. improvements of this type of inhibitors have been made by exchanging the morpholines with bridged morpholines in pyrazolopyrimidine