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Inhibition of mutant EGFR in lung cancer cells triggers SOX2-FOXO6-dependent survival pathways.

Rothenberg SM, Concannon K, Cullen S, Boulay G, Turke AB, Faber AC, Lockerman EL, Rivera MN, Engelman JA, Maheswaran S, Haber DA - Elife (2015)

Bottom Line: Treatment of EGFR-mutant lung cancer with erlotinib results in dramatic tumor regression but it is invariably followed by drug resistance.In characterizing early transcriptional changes following drug treatment of mutant EGFR-addicted cells, we identified the stem cell transcriptional regulator SOX2 as being rapidly and specifically induced, both in vitro and in vivo.Together, these observations point to a physiological feedback mechanism that attenuates oncogene addiction-mediated cell death associated with the withdrawal of growth factor signaling and may therefore contribute to the development of resistance.

View Article: PubMed Central - PubMed

Affiliation: Cancer Center, Massachusetts General Hospital, Harvard Medical School, Charlestown, United States.

ABSTRACT
Treatment of EGFR-mutant lung cancer with erlotinib results in dramatic tumor regression but it is invariably followed by drug resistance. In characterizing early transcriptional changes following drug treatment of mutant EGFR-addicted cells, we identified the stem cell transcriptional regulator SOX2 as being rapidly and specifically induced, both in vitro and in vivo. Suppression of SOX2 sensitizes cells to erlotinib-mediated apoptosis, ultimately decreasing the emergence of acquired resistance, whereas its ectopic expression reduces drug-induced cell death. We show that erlotinib relieves EGFR-dependent suppression of FOXO6, leading to its induction of SOX2, which in turn represses the pro-apoptotic BH3-only genes BIM and BMF. Together, these observations point to a physiological feedback mechanism that attenuates oncogene addiction-mediated cell death associated with the withdrawal of growth factor signaling and may therefore contribute to the development of resistance.

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Stem cell markers do not colocalize with SOX2+ cells.Immunofluorescence microscopy was carried out on erlotinib-treated HCC827(upper panels) or PC9 (lower panels) cells using antibodies to SOX2 andvarious stem cell markers. CD133 (upper row of panels) could be detectedin a rare population of PC9 cells (but not HCC827 cells) that was clearlymutually exclusive from SOX2+ cells, while CD44 and MYC wereexpressed in the majority of cells, irrespective of SOX2 expression.Neither membrane localization of CD24 (rightmost panel, middle rows) norexpression of OCT4 and KLF4 (data not shown) could be detected in eithercell line.DOI:http://dx.doi.org/10.7554/eLife.06132.016
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fig2s2: Stem cell markers do not colocalize with SOX2+ cells.Immunofluorescence microscopy was carried out on erlotinib-treated HCC827(upper panels) or PC9 (lower panels) cells using antibodies to SOX2 andvarious stem cell markers. CD133 (upper row of panels) could be detectedin a rare population of PC9 cells (but not HCC827 cells) that was clearlymutually exclusive from SOX2+ cells, while CD44 and MYC wereexpressed in the majority of cells, irrespective of SOX2 expression.Neither membrane localization of CD24 (rightmost panel, middle rows) norexpression of OCT4 and KLF4 (data not shown) could be detected in eithercell line.DOI:http://dx.doi.org/10.7554/eLife.06132.016

Mentions: The level of SOX2 induction in cultured cells exposed to erlotinib showedconsiderable heterogeneity, with a subset of cells (∼20%, with someexperimental variability) expressing high levels (Figure 2A). The SOX2+ fraction was not increased by higher drugdosage, beyond that required for full inhibition of EGFR (Figure 2—figure supplement 1). Given the link betweenSOX2 expression and cellular reprogramming, we first asked whether cells with thehigh SOX2 expression represent a subset with stem cell markers. However, SOX2expression did not correlate with expression of the putative stem cell markers CD133,CD44, CD24, OCT-4, or KLF-4 (Figure 2—figuresupplement 2) nor did microarray-based expression profiling of highSOX2-sorted cells identify a stem-like signature (data not shown). Nonetheless,SOX2-expressing cells had a very low proliferative index, as measured by Ki67staining (0.5% of Ki67+/SOX2+ vs 51% Ki67+/SOX2− HCC827cells at baseline [p = 0.015]; and 0.15% Ki67+/SOX2+ vs 6.4%Ki67+/SOX2− cells following erlotinib [p < 0.0001]) (Figure 2C, Figure 2—figure supplement 3).10.7554/eLife.06132.008Figure 2.Induction of SOX2 in erlotinib-treated cells.


Inhibition of mutant EGFR in lung cancer cells triggers SOX2-FOXO6-dependent survival pathways.

Rothenberg SM, Concannon K, Cullen S, Boulay G, Turke AB, Faber AC, Lockerman EL, Rivera MN, Engelman JA, Maheswaran S, Haber DA - Elife (2015)

Stem cell markers do not colocalize with SOX2+ cells.Immunofluorescence microscopy was carried out on erlotinib-treated HCC827(upper panels) or PC9 (lower panels) cells using antibodies to SOX2 andvarious stem cell markers. CD133 (upper row of panels) could be detectedin a rare population of PC9 cells (but not HCC827 cells) that was clearlymutually exclusive from SOX2+ cells, while CD44 and MYC wereexpressed in the majority of cells, irrespective of SOX2 expression.Neither membrane localization of CD24 (rightmost panel, middle rows) norexpression of OCT4 and KLF4 (data not shown) could be detected in eithercell line.DOI:http://dx.doi.org/10.7554/eLife.06132.016
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4384750&req=5

fig2s2: Stem cell markers do not colocalize with SOX2+ cells.Immunofluorescence microscopy was carried out on erlotinib-treated HCC827(upper panels) or PC9 (lower panels) cells using antibodies to SOX2 andvarious stem cell markers. CD133 (upper row of panels) could be detectedin a rare population of PC9 cells (but not HCC827 cells) that was clearlymutually exclusive from SOX2+ cells, while CD44 and MYC wereexpressed in the majority of cells, irrespective of SOX2 expression.Neither membrane localization of CD24 (rightmost panel, middle rows) norexpression of OCT4 and KLF4 (data not shown) could be detected in eithercell line.DOI:http://dx.doi.org/10.7554/eLife.06132.016
Mentions: The level of SOX2 induction in cultured cells exposed to erlotinib showedconsiderable heterogeneity, with a subset of cells (∼20%, with someexperimental variability) expressing high levels (Figure 2A). The SOX2+ fraction was not increased by higher drugdosage, beyond that required for full inhibition of EGFR (Figure 2—figure supplement 1). Given the link betweenSOX2 expression and cellular reprogramming, we first asked whether cells with thehigh SOX2 expression represent a subset with stem cell markers. However, SOX2expression did not correlate with expression of the putative stem cell markers CD133,CD44, CD24, OCT-4, or KLF-4 (Figure 2—figuresupplement 2) nor did microarray-based expression profiling of highSOX2-sorted cells identify a stem-like signature (data not shown). Nonetheless,SOX2-expressing cells had a very low proliferative index, as measured by Ki67staining (0.5% of Ki67+/SOX2+ vs 51% Ki67+/SOX2− HCC827cells at baseline [p = 0.015]; and 0.15% Ki67+/SOX2+ vs 6.4%Ki67+/SOX2− cells following erlotinib [p < 0.0001]) (Figure 2C, Figure 2—figure supplement 3).10.7554/eLife.06132.008Figure 2.Induction of SOX2 in erlotinib-treated cells.

Bottom Line: Treatment of EGFR-mutant lung cancer with erlotinib results in dramatic tumor regression but it is invariably followed by drug resistance.In characterizing early transcriptional changes following drug treatment of mutant EGFR-addicted cells, we identified the stem cell transcriptional regulator SOX2 as being rapidly and specifically induced, both in vitro and in vivo.Together, these observations point to a physiological feedback mechanism that attenuates oncogene addiction-mediated cell death associated with the withdrawal of growth factor signaling and may therefore contribute to the development of resistance.

View Article: PubMed Central - PubMed

Affiliation: Cancer Center, Massachusetts General Hospital, Harvard Medical School, Charlestown, United States.

ABSTRACT
Treatment of EGFR-mutant lung cancer with erlotinib results in dramatic tumor regression but it is invariably followed by drug resistance. In characterizing early transcriptional changes following drug treatment of mutant EGFR-addicted cells, we identified the stem cell transcriptional regulator SOX2 as being rapidly and specifically induced, both in vitro and in vivo. Suppression of SOX2 sensitizes cells to erlotinib-mediated apoptosis, ultimately decreasing the emergence of acquired resistance, whereas its ectopic expression reduces drug-induced cell death. We show that erlotinib relieves EGFR-dependent suppression of FOXO6, leading to its induction of SOX2, which in turn represses the pro-apoptotic BH3-only genes BIM and BMF. Together, these observations point to a physiological feedback mechanism that attenuates oncogene addiction-mediated cell death associated with the withdrawal of growth factor signaling and may therefore contribute to the development of resistance.

Show MeSH
Related in: MedlinePlus