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Inhibition of SHP2 in basal-like and triple-negative breast cells induces basal-to-luminal transition, hormone dependency, and sensitivity to anti-hormone treatment.

Zhao H, Agazie YM - BMC Cancer (2015)

Bottom Line: The Src homology phosphotyrosyl phosphatase 2 (SHP2) is a positive effector of cell growth and survival signaling as well transformation induced by multiple tyrosine kinase oncogenes.The occurrence of BLT was confirmed by the loss of the basal marker alpha smooth muscle actin and the acquisition of the luminal marker cytokeratin 18 (CK18) expression.Our data show that inhibition of SHP2 induces BLT, ERα expression, dependency on estrogen for growth, and sensitivity to anti-hormone therapy.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and The Marry Babb Randolph Cancer Center School of Medicine, West Virginia University, Morgantown, WV, 26506, USA. ttangela168@gmail.com.

ABSTRACT

Background: The Src homology phosphotyrosyl phosphatase 2 (SHP2) is a positive effector of cell growth and survival signaling as well transformation induced by multiple tyrosine kinase oncogenes. Since the basal-like and triple-negative breast cancer (BTBC) is characterized by dysregulation of multiple tyrosine kinase oncogenes, we wanted to determine the importance of SHP2 in BTBC cell lines.

Methods: Short hairpin RNA-based and dominant-negative expression-based SHP2 inhibition techniques were used to interrogate the functional importance of SHP2 in BTBC cell biology. In addition, cell viability and proliferation assays were used to determine hormone dependency for growth and sensitivity to anti-estrogen treatment.

Results: We show that inhibition of SHP2 in BTBC cells induces luminal-like epithelial morphology while suppressing the mesenchymal and invasive property. We have termed this process as basal-to-luminal transition (BLT). The occurrence of BLT was confirmed by the loss of the basal marker alpha smooth muscle actin and the acquisition of the luminal marker cytokeratin 18 (CK18) expression. Furthermore, the occurrence of BLT led to estrogen receptor alpha (ERα) expression, hormone dependency, and sensitivity to tamoxifen treatment.

Conclusions: Our data show that inhibition of SHP2 induces BLT, ERα expression, dependency on estrogen for growth, and sensitivity to anti-hormone therapy. Therefore, inhibition of SHP2 may provide a therapeutic benefit in basal-like and triple-negative breast cancer.

No MeSH data available.


Related in: MedlinePlus

Effect of SHP2 silencing on estrogen responsiveness. A) Comparison of cells for responsiveness to estradiol treatment using cell viability assay. B) Effect of estradiol on the growth of the control and the SHP2-silenced MDA-MB231 and MDA-MB468 cells as determined by direct cell counting in a time-course fashion. C) Effect of SHP2 silencing on sensitivity to tamoxifen in the MDA-MB231 cells. D) Effect on SHP2 silencing on sensitivity to tamoxifen in the MDA-MB468 cells. V: vehicle; E2: estradiol; Tam: tamoxifen; Con: control shRNA; sh: SHP2 shRNA.
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Fig5: Effect of SHP2 silencing on estrogen responsiveness. A) Comparison of cells for responsiveness to estradiol treatment using cell viability assay. B) Effect of estradiol on the growth of the control and the SHP2-silenced MDA-MB231 and MDA-MB468 cells as determined by direct cell counting in a time-course fashion. C) Effect of SHP2 silencing on sensitivity to tamoxifen in the MDA-MB231 cells. D) Effect on SHP2 silencing on sensitivity to tamoxifen in the MDA-MB468 cells. V: vehicle; E2: estradiol; Tam: tamoxifen; Con: control shRNA; sh: SHP2 shRNA.

Mentions: The expression of ERα (Figure 4C) in SHP2-silenced cells suggested that they may depend on estrogen signaling for growth. To test this possibility, we conducted cell growth studies in the presence and absence of estradiol (E2) using a cell viability assay (Promega) that determines cell growth based on ATP levels. The MCF-10A (non-transformed breast epithelial cells) and the MCF-7 (the hormone-positive breast cancer cell line) cells that require E2 for growth were used as positive controls. While the control cells did not respond to E2 treatment, the SHP2-silenced cells showed an approximately 30% increase in cell growth rate over the unstimulated counterparts (Figure 5A). As expected, the growth of the MCF-10A and the MCF-7 cells was retarded by E2 subtraction in the growth medium. We also determined the effect of E2 on cell growth rate by direct counting in a time-course fashion as described previously [32]. Consistent with the cell viability data, addition of E2 in the growth medium increased the growth rate of the SHP2-slinced cells. For instance, the unstimulated and the E2-stimulated SHP2-silenced cells grew by approximately 4-fold and 7-fold, respectively, over a period of 3 days (Figure 5B). As expected, the control cells grew by more than 10-fold over the same period of time. These observations suggest that SHP2 inhibition-induced ERα expression confers hormone dependency.Figure 5


Inhibition of SHP2 in basal-like and triple-negative breast cells induces basal-to-luminal transition, hormone dependency, and sensitivity to anti-hormone treatment.

Zhao H, Agazie YM - BMC Cancer (2015)

Effect of SHP2 silencing on estrogen responsiveness. A) Comparison of cells for responsiveness to estradiol treatment using cell viability assay. B) Effect of estradiol on the growth of the control and the SHP2-silenced MDA-MB231 and MDA-MB468 cells as determined by direct cell counting in a time-course fashion. C) Effect of SHP2 silencing on sensitivity to tamoxifen in the MDA-MB231 cells. D) Effect on SHP2 silencing on sensitivity to tamoxifen in the MDA-MB468 cells. V: vehicle; E2: estradiol; Tam: tamoxifen; Con: control shRNA; sh: SHP2 shRNA.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4359540&req=5

Fig5: Effect of SHP2 silencing on estrogen responsiveness. A) Comparison of cells for responsiveness to estradiol treatment using cell viability assay. B) Effect of estradiol on the growth of the control and the SHP2-silenced MDA-MB231 and MDA-MB468 cells as determined by direct cell counting in a time-course fashion. C) Effect of SHP2 silencing on sensitivity to tamoxifen in the MDA-MB231 cells. D) Effect on SHP2 silencing on sensitivity to tamoxifen in the MDA-MB468 cells. V: vehicle; E2: estradiol; Tam: tamoxifen; Con: control shRNA; sh: SHP2 shRNA.
Mentions: The expression of ERα (Figure 4C) in SHP2-silenced cells suggested that they may depend on estrogen signaling for growth. To test this possibility, we conducted cell growth studies in the presence and absence of estradiol (E2) using a cell viability assay (Promega) that determines cell growth based on ATP levels. The MCF-10A (non-transformed breast epithelial cells) and the MCF-7 (the hormone-positive breast cancer cell line) cells that require E2 for growth were used as positive controls. While the control cells did not respond to E2 treatment, the SHP2-silenced cells showed an approximately 30% increase in cell growth rate over the unstimulated counterparts (Figure 5A). As expected, the growth of the MCF-10A and the MCF-7 cells was retarded by E2 subtraction in the growth medium. We also determined the effect of E2 on cell growth rate by direct counting in a time-course fashion as described previously [32]. Consistent with the cell viability data, addition of E2 in the growth medium increased the growth rate of the SHP2-slinced cells. For instance, the unstimulated and the E2-stimulated SHP2-silenced cells grew by approximately 4-fold and 7-fold, respectively, over a period of 3 days (Figure 5B). As expected, the control cells grew by more than 10-fold over the same period of time. These observations suggest that SHP2 inhibition-induced ERα expression confers hormone dependency.Figure 5

Bottom Line: The Src homology phosphotyrosyl phosphatase 2 (SHP2) is a positive effector of cell growth and survival signaling as well transformation induced by multiple tyrosine kinase oncogenes.The occurrence of BLT was confirmed by the loss of the basal marker alpha smooth muscle actin and the acquisition of the luminal marker cytokeratin 18 (CK18) expression.Our data show that inhibition of SHP2 induces BLT, ERα expression, dependency on estrogen for growth, and sensitivity to anti-hormone therapy.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and The Marry Babb Randolph Cancer Center School of Medicine, West Virginia University, Morgantown, WV, 26506, USA. ttangela168@gmail.com.

ABSTRACT

Background: The Src homology phosphotyrosyl phosphatase 2 (SHP2) is a positive effector of cell growth and survival signaling as well transformation induced by multiple tyrosine kinase oncogenes. Since the basal-like and triple-negative breast cancer (BTBC) is characterized by dysregulation of multiple tyrosine kinase oncogenes, we wanted to determine the importance of SHP2 in BTBC cell lines.

Methods: Short hairpin RNA-based and dominant-negative expression-based SHP2 inhibition techniques were used to interrogate the functional importance of SHP2 in BTBC cell biology. In addition, cell viability and proliferation assays were used to determine hormone dependency for growth and sensitivity to anti-estrogen treatment.

Results: We show that inhibition of SHP2 in BTBC cells induces luminal-like epithelial morphology while suppressing the mesenchymal and invasive property. We have termed this process as basal-to-luminal transition (BLT). The occurrence of BLT was confirmed by the loss of the basal marker alpha smooth muscle actin and the acquisition of the luminal marker cytokeratin 18 (CK18) expression. Furthermore, the occurrence of BLT led to estrogen receptor alpha (ERα) expression, hormone dependency, and sensitivity to tamoxifen treatment.

Conclusions: Our data show that inhibition of SHP2 induces BLT, ERα expression, dependency on estrogen for growth, and sensitivity to anti-hormone therapy. Therefore, inhibition of SHP2 may provide a therapeutic benefit in basal-like and triple-negative breast cancer.

No MeSH data available.


Related in: MedlinePlus