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. 2011;6(9):e24849.
doi: 10.1371/journal.pone.0024849. Epub 2011 Sep 16.

Simultaneous inhibition of mTOR-containing complex 1 (mTORC1) and MNK induces apoptosis of cutaneous T-cell lymphoma (CTCL) cells

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Simultaneous inhibition of mTOR-containing complex 1 (mTORC1) and MNK induces apoptosis of cutaneous T-cell lymphoma (CTCL) cells

Michal Marzec et al. PLoS One. 2011.

Retraction in

Abstract

Background: mTOR kinase forms the mTORC1 complex by associating with raptor and other proteins and affects a number of key cell functions. mTORC1 activates p70S6kinase 1 (p70S6K1) and inhibits 4E-binding protein 1 (4E-BP1). In turn, p70S6K1 phosphorylates a S6 protein of the 40S ribosomal subunit (S6rp) and 4E-BP1, with the latter negatively regulating eukaryotic initiation factor 4E (eIF-4E). MNK1 and MNK2 kinases phosphorylate and augment activity of eIF4E. Rapamycin and its analogs are highly specific, potent, and relatively non-toxic inhibitors of mTORC1. Although mTORC1 activation is present in many types of malignancies, rapamycin-type inhibitors shows relatively limited clinical efficacy as single agents. Initially usually indolent, CTCL displays a tendency to progress to the aggressive forms with limited response to therapy and poor prognosis. Our previous study (M. Marzec et al. 2008) has demonstrated that CTCL cells display mTORC1 activation and short-term treatment of CTCL-derived cells with rapamycin suppressed their proliferation and had little effect on the cell survival.

Methods: Cells derived from CTCL were treated with mTORC1 inhibitor rapamycin and MNK inhibitor and evaluated for inhibition of the mTORC1 signaling pathway and cell growth and survival.

Results: Whereas the treatment with rapamycin persistently inhibited mTORC1 signaling, it suppressed only partially the cell growth. MNK kinase mediated the eIF4E phosphorylation and inhibition or depletion of MNK markedly suppressed proliferation of the CTCL cells when combined with the rapamycin-mediated inhibition of mTORC1. While MNK inhibition alone mildly suppressed the CTCL cell growth, the combined MNK and mTORC1 inhibition totally abrogated the growth. Similarly, MNK inhibitor alone displayed a minimal pro-apoptotic effect; in combination with rapamycin it triggered profound cell apoptosis.

Conclusions: These findings indicate that the combined inhibition of mTORC1 and MNK may prove beneficial in the treatment of CTCL and other malignancies.

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Conflict of interest statement

Competing Interests: The authors have declared that no competing interests exist.

Figures

Figure 1
Figure 1. Rapamycin partially inhibits proliferation of T-cell lymphoma cells and promotes eIF4E S209 phosphorylation.
The CTCL-derived cell lines MyLa2059 and MyLa3675 were treated in triplicates with 200 nM of rapamycin (mTORC1i) for up to 10 days and (A) labeled with BrdU for the last 4 hr of the culture and examined using the EIA plate reader (the result shows mean value of four separate experiments) or (B) lysed and analyzed by Western blotting with antibodies against the depicted phosphorylated and total proteins with detection of actin serving as the loading control.
Figure 2
Figure 2. MNK inhibition suppresses eIF4E S209 phosphorylation.
A: effect of MNK1/2 inhibitor (MNKi) used at various concentrations on eIF4E S209 phosphorylation in MyLa2059 and MyLa3675 cell lines with Akt T308 phosphorylation serving as a control. B: effect of simultaneous application of MNKi (5 µM) and mTORC1i (200 nM) on phosphorylation of eIF4E S209, S6rp S235/236, and MNK T197/202 and expression of the anti-apoptotic proteins Mcl-1 and BcL-xL. C: effect of siRNA-mediated MNK1 and MNK2 depletion on eIF4E S209 phosphorylation in the presence or absence of rapamycin. D: effect of simultaneous exposure of IL-2-dependent CTCL cell line Sez-4 to MNKi and mTORC1i on phosphorylation and expression the depicted proteins. E: effect of simultaneous treatment of patient-derived, primed Sezary CTCL cells to MNKi and mTORC1i on phosphorylation and expression the depicted proteins.
Figure 3
Figure 3. Lack of effect on eIF4E phosphorylation of several kinase inhibitors.
A and B: Expression of eIF4E S209 was examined after treatment of MyLa 2059 cells with inhibitor of p38 MAPK, MEK1/2 (U0126), PKC (Go6983), PKA (Bisindolylmaleimide I) and PI3K (Wortmannin and LY294002). Expression of phospho-Akt T308 and -ERK1/2 T202/Y204 and actin and cell treatment with MNKi served as controls.
Figure 4
Figure 4. Simultaneous mTORC1 and MNK inhibition suppresses growth and induces apoptosis of CTCL-derived cells.

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