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GSK3 is required for rapalogs to induce degradation of some oncogenic proteins and to suppress cancer cell growth.

Koo J, Wang X, Owonikoko TK, Ramalingam SS, Khuri FR, Sun SY - Oncotarget (2015)

Bottom Line: GSK3 inhibition rescued rapamcyin-induced reduction of several oncogenic proteins such as cyclin D1, Mcl-1 and c-Myc, without interfering with the ability of rapamycin to suppress mTORC1 signaling and cap binding.Interestingly, rapamycin induces proteasomal degradation of these oncogenic proteins, as evidenced by their decreased stabilities induced by rapamcyin and rescue of their reduction by proteasomal inhibition.Thus, induction of GSK3-dependent degradation of these oncogenic proteins is likely secondary to mTORC2 inhibition; this effect should be critical for rapamycin to exert its anticancer activity.

View Article: PubMed Central - PubMed

Affiliation: Department of Hematology and Medical Oncology, Emory University School of Medicine and Winship Cancer Institute, Atlanta, GA, USA.

ABSTRACT
The single-agent activity of rapalogs (rapamycin and its analogues) in most tumor types has been modest at best. The underlying mechanisms are largely unclear. In this report, we have uncovered a critical role of GSK3 in regulating degradation of some oncogenic proteins induced by rapalogs and cell sensitivity to rapalogs. The basal level of GSK3 activity was positively correlated with cell sensitivity of lung cancer cell lines to rapalogs. GSK3 inhibition antagonized rapamycin's growth inhibitory effects both in vitro and in vivo, while enforced activation of GSK3β sensitized cells to rapamycin. GSK3 inhibition rescued rapamcyin-induced reduction of several oncogenic proteins such as cyclin D1, Mcl-1 and c-Myc, without interfering with the ability of rapamycin to suppress mTORC1 signaling and cap binding. Interestingly, rapamycin induces proteasomal degradation of these oncogenic proteins, as evidenced by their decreased stabilities induced by rapamcyin and rescue of their reduction by proteasomal inhibition. Moreover, acute or short-time rapamycin treatment dissociated not only raptor, but also rictor from mTOR in several tested cell lines, suggesting inhibition of both mTORC1 and mTORC2. Thus, induction of GSK3-dependent degradation of these oncogenic proteins is likely secondary to mTORC2 inhibition; this effect should be critical for rapamycin to exert its anticancer activity.

No MeSH data available.


Related in: MedlinePlus

Basal levels of p-GSK3 in human lung cancer cell lines (A) are inversely correlated with cell sensitivity to rapalogs (B and C)Whole-cell lysates were prepared from the listed cell lines with comparable cell densities and subjected to Western blotting for detection of the indicated proteins (A). The intensities of these proteins were quantified with NIH Image J software. The growth-inhibitory effects of rapamycin or RAD001 at 10 nM were determined with the SRB assay after 3 days. The correlation between p-GSK3/GSK3 and growth inhibition was calculated with GraphPad InStat software (B and C).
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Figure 3: Basal levels of p-GSK3 in human lung cancer cell lines (A) are inversely correlated with cell sensitivity to rapalogs (B and C)Whole-cell lysates were prepared from the listed cell lines with comparable cell densities and subjected to Western blotting for detection of the indicated proteins (A). The intensities of these proteins were quantified with NIH Image J software. The growth-inhibitory effects of rapamycin or RAD001 at 10 nM were determined with the SRB assay after 3 days. The correlation between p-GSK3/GSK3 and growth inhibition was calculated with GraphPad InStat software (B and C).

Mentions: Following these findings, we further determined whether basal levels of GSK3 activity are associated with cell sensitivity to rapalogs. Here we detected basal levels of p-GSK3 as an indication of inactivated or low GSK3 activity in a panel of 13 NSCLC cell lines by Western blotting (Fig. 3A). These cell lines possessed different sensitivities to rapamycin or RAD001 as determined in a 3-day growth-inhibitory assay (Fig. 3B). Correlation analysis showed that high p-GSK3 levels (i.e., low GSK3 activity) were significantly associated with weak growth-inhibitory effect of rapamycin (r = 0.7895, P = 0.0013) or RAD001 (r = 0.7870, P = 0.0014) (Fig. 3C), meaning that low GSK3 activity is associated with reduced cell sensitivity to rapalogs (e.g., in NSCLC cell lines).


GSK3 is required for rapalogs to induce degradation of some oncogenic proteins and to suppress cancer cell growth.

Koo J, Wang X, Owonikoko TK, Ramalingam SS, Khuri FR, Sun SY - Oncotarget (2015)

Basal levels of p-GSK3 in human lung cancer cell lines (A) are inversely correlated with cell sensitivity to rapalogs (B and C)Whole-cell lysates were prepared from the listed cell lines with comparable cell densities and subjected to Western blotting for detection of the indicated proteins (A). The intensities of these proteins were quantified with NIH Image J software. The growth-inhibitory effects of rapamycin or RAD001 at 10 nM were determined with the SRB assay after 3 days. The correlation between p-GSK3/GSK3 and growth inhibition was calculated with GraphPad InStat software (B and C).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4496196&req=5

Figure 3: Basal levels of p-GSK3 in human lung cancer cell lines (A) are inversely correlated with cell sensitivity to rapalogs (B and C)Whole-cell lysates were prepared from the listed cell lines with comparable cell densities and subjected to Western blotting for detection of the indicated proteins (A). The intensities of these proteins were quantified with NIH Image J software. The growth-inhibitory effects of rapamycin or RAD001 at 10 nM were determined with the SRB assay after 3 days. The correlation between p-GSK3/GSK3 and growth inhibition was calculated with GraphPad InStat software (B and C).
Mentions: Following these findings, we further determined whether basal levels of GSK3 activity are associated with cell sensitivity to rapalogs. Here we detected basal levels of p-GSK3 as an indication of inactivated or low GSK3 activity in a panel of 13 NSCLC cell lines by Western blotting (Fig. 3A). These cell lines possessed different sensitivities to rapamycin or RAD001 as determined in a 3-day growth-inhibitory assay (Fig. 3B). Correlation analysis showed that high p-GSK3 levels (i.e., low GSK3 activity) were significantly associated with weak growth-inhibitory effect of rapamycin (r = 0.7895, P = 0.0013) or RAD001 (r = 0.7870, P = 0.0014) (Fig. 3C), meaning that low GSK3 activity is associated with reduced cell sensitivity to rapalogs (e.g., in NSCLC cell lines).

Bottom Line: GSK3 inhibition rescued rapamcyin-induced reduction of several oncogenic proteins such as cyclin D1, Mcl-1 and c-Myc, without interfering with the ability of rapamycin to suppress mTORC1 signaling and cap binding.Interestingly, rapamycin induces proteasomal degradation of these oncogenic proteins, as evidenced by their decreased stabilities induced by rapamcyin and rescue of their reduction by proteasomal inhibition.Thus, induction of GSK3-dependent degradation of these oncogenic proteins is likely secondary to mTORC2 inhibition; this effect should be critical for rapamycin to exert its anticancer activity.

View Article: PubMed Central - PubMed

Affiliation: Department of Hematology and Medical Oncology, Emory University School of Medicine and Winship Cancer Institute, Atlanta, GA, USA.

ABSTRACT
The single-agent activity of rapalogs (rapamycin and its analogues) in most tumor types has been modest at best. The underlying mechanisms are largely unclear. In this report, we have uncovered a critical role of GSK3 in regulating degradation of some oncogenic proteins induced by rapalogs and cell sensitivity to rapalogs. The basal level of GSK3 activity was positively correlated with cell sensitivity of lung cancer cell lines to rapalogs. GSK3 inhibition antagonized rapamycin's growth inhibitory effects both in vitro and in vivo, while enforced activation of GSK3β sensitized cells to rapamycin. GSK3 inhibition rescued rapamcyin-induced reduction of several oncogenic proteins such as cyclin D1, Mcl-1 and c-Myc, without interfering with the ability of rapamycin to suppress mTORC1 signaling and cap binding. Interestingly, rapamycin induces proteasomal degradation of these oncogenic proteins, as evidenced by their decreased stabilities induced by rapamcyin and rescue of their reduction by proteasomal inhibition. Moreover, acute or short-time rapamycin treatment dissociated not only raptor, but also rictor from mTOR in several tested cell lines, suggesting inhibition of both mTORC1 and mTORC2. Thus, induction of GSK3-dependent degradation of these oncogenic proteins is likely secondary to mTORC2 inhibition; this effect should be critical for rapamycin to exert its anticancer activity.

No MeSH data available.


Related in: MedlinePlus