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Triple negative breast cancers comprise a highly tumorigenic cell subpopulation detectable by its high responsiveness to a Sox2 regulatory region 2 (SRR2) reporter.

Jung K, Gupta N, Wang P, Lewis JT, Gopal K, Wu F, Ye X, Alshareef A, Abdulkarim BS, Douglas DN, Kneteman NM, Lai R - Oncotarget (2015)

Bottom Line: We have recently described a novel phenotypic dichotomy within estrogen receptor-positive breast cancer cells; the cell subset responsive to a Sox2 regulatory region (SRR2) reporter (RR cells) are significantly more tumorigenic than the reporter unresponsive (RU) cells.Third, within the CD44(High)/CD24- tumor-initiating cell population derived from MDA-MB-231, RR cells were significantly more tumorigenic than RU cells in an in vivo SCID/Beige xenograft mouse model.To conclude, we described a novel phenotypic heterogeneity within TNBC, and the SRR2 reporter responsiveness is a useful marker for identifying a highly tumorigenic cell subset within the CD44(High)/CD24-tumor-initiating cell population.

View Article: PubMed Central - PubMed

Affiliation: Department of Oncology, University of Alberta, Edmonton, Alberta, Canada.

ABSTRACT
We have recently described a novel phenotypic dichotomy within estrogen receptor-positive breast cancer cells; the cell subset responsive to a Sox2 regulatory region (SRR2) reporter (RR cells) are significantly more tumorigenic than the reporter unresponsive (RU) cells. Here, we report that a similar phenomenon also exists in triple negative breast cancer (TNBC), with RR cells more tumorigenic than RU cells. First, examination of all 3 TNBC cell lines stably infected with the SRR2 reporter revealed the presence of a cell subset exhibiting reporter activity. Second, RU and RR cells purified by flow cytometry showed that RR cells expressed higher levels of CD44, generated more spheres in a limiting dilution mammosphere formation assay, and formed larger and more complex structures in Matrigel. Third, within the CD44(High)/CD24- tumor-initiating cell population derived from MDA-MB-231, RR cells were significantly more tumorigenic than RU cells in an in vivo SCID/Beige xenograft mouse model. Examination of 4 TNBC tumors from patients also revealed the presence of a RR cell subset, ranging from 1.1-3.8%. To conclude, we described a novel phenotypic heterogeneity within TNBC, and the SRR2 reporter responsiveness is a useful marker for identifying a highly tumorigenic cell subset within the CD44(High)/CD24-tumor-initiating cell population.

No MeSH data available.


Related in: MedlinePlus

TNBC cell lines comprise of cells with heterogeneous Sox2 regulatory region 2 (SRR2) reporter activityA. FACS dot plots illustrating the GFP expression of ER− cell lines virally-infected with the mCMV or SRR2 reporter plasmids. Gates drawn show the RU and RR subsets collected and cultured separately thereafter, percent of gated live population is reported. B. Flow cytometry dot plot and merged histogram analyses for GFP expression of ER− RU and RR lines. Cells were also harvested and assayed for relative SRR2 luciferase activity. C. Q-PCR results of SOX2 and OCT4A expression in the triple-negative RU and RR cell lines normalized to GAPDH, and further normalized to MCF7 RU sample. Previously reported high Sox2-expressing MCF7 RU and RR cell lines SOX2 and OCT4A expression data are shown for comparison. Western blot visualizing Sox2 and Oct4A/B protein expression. Ntera2 (a malignant human pluripotent embryonic carcinoma cell line) acts a positive control for Sox2 and Oct4A/B expression.
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Figure 2: TNBC cell lines comprise of cells with heterogeneous Sox2 regulatory region 2 (SRR2) reporter activityA. FACS dot plots illustrating the GFP expression of ER− cell lines virally-infected with the mCMV or SRR2 reporter plasmids. Gates drawn show the RU and RR subsets collected and cultured separately thereafter, percent of gated live population is reported. B. Flow cytometry dot plot and merged histogram analyses for GFP expression of ER− RU and RR lines. Cells were also harvested and assayed for relative SRR2 luciferase activity. C. Q-PCR results of SOX2 and OCT4A expression in the triple-negative RU and RR cell lines normalized to GAPDH, and further normalized to MCF7 RU sample. Previously reported high Sox2-expressing MCF7 RU and RR cell lines SOX2 and OCT4A expression data are shown for comparison. Western blot visualizing Sox2 and Oct4A/B protein expression. Ntera2 (a malignant human pluripotent embryonic carcinoma cell line) acts a positive control for Sox2 and Oct4A/B expression.

Mentions: To facilitate our studies, we established TNBC cell clones stably transfected with the SRR2 reporter using a lentiviral infection protocol described previously [7]. Three TNBC cell lines, including MDA-MB-231, MDA-MB-468 and SUM-149, were included for this study. Cells from these three cell lines stably transfected with the minimal CMV reporter served as the negative controls. To detect evidence of responsiveness to the SRR2 reporter, we performed flow cytometry to detect GFP expression. At two weeks after the lentiviral infection, all three cell lines showed reporter responsiveness in a subset of cells, with 34.3% in MDA-MB-231, 16.3% in MDA-MB-468 and 48.9% in SUM149, as compared to the mCMV reporter cells (Figure 2A).


Triple negative breast cancers comprise a highly tumorigenic cell subpopulation detectable by its high responsiveness to a Sox2 regulatory region 2 (SRR2) reporter.

Jung K, Gupta N, Wang P, Lewis JT, Gopal K, Wu F, Ye X, Alshareef A, Abdulkarim BS, Douglas DN, Kneteman NM, Lai R - Oncotarget (2015)

TNBC cell lines comprise of cells with heterogeneous Sox2 regulatory region 2 (SRR2) reporter activityA. FACS dot plots illustrating the GFP expression of ER− cell lines virally-infected with the mCMV or SRR2 reporter plasmids. Gates drawn show the RU and RR subsets collected and cultured separately thereafter, percent of gated live population is reported. B. Flow cytometry dot plot and merged histogram analyses for GFP expression of ER− RU and RR lines. Cells were also harvested and assayed for relative SRR2 luciferase activity. C. Q-PCR results of SOX2 and OCT4A expression in the triple-negative RU and RR cell lines normalized to GAPDH, and further normalized to MCF7 RU sample. Previously reported high Sox2-expressing MCF7 RU and RR cell lines SOX2 and OCT4A expression data are shown for comparison. Western blot visualizing Sox2 and Oct4A/B protein expression. Ntera2 (a malignant human pluripotent embryonic carcinoma cell line) acts a positive control for Sox2 and Oct4A/B expression.
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Related In: Results  -  Collection

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Figure 2: TNBC cell lines comprise of cells with heterogeneous Sox2 regulatory region 2 (SRR2) reporter activityA. FACS dot plots illustrating the GFP expression of ER− cell lines virally-infected with the mCMV or SRR2 reporter plasmids. Gates drawn show the RU and RR subsets collected and cultured separately thereafter, percent of gated live population is reported. B. Flow cytometry dot plot and merged histogram analyses for GFP expression of ER− RU and RR lines. Cells were also harvested and assayed for relative SRR2 luciferase activity. C. Q-PCR results of SOX2 and OCT4A expression in the triple-negative RU and RR cell lines normalized to GAPDH, and further normalized to MCF7 RU sample. Previously reported high Sox2-expressing MCF7 RU and RR cell lines SOX2 and OCT4A expression data are shown for comparison. Western blot visualizing Sox2 and Oct4A/B protein expression. Ntera2 (a malignant human pluripotent embryonic carcinoma cell line) acts a positive control for Sox2 and Oct4A/B expression.
Mentions: To facilitate our studies, we established TNBC cell clones stably transfected with the SRR2 reporter using a lentiviral infection protocol described previously [7]. Three TNBC cell lines, including MDA-MB-231, MDA-MB-468 and SUM-149, were included for this study. Cells from these three cell lines stably transfected with the minimal CMV reporter served as the negative controls. To detect evidence of responsiveness to the SRR2 reporter, we performed flow cytometry to detect GFP expression. At two weeks after the lentiviral infection, all three cell lines showed reporter responsiveness in a subset of cells, with 34.3% in MDA-MB-231, 16.3% in MDA-MB-468 and 48.9% in SUM149, as compared to the mCMV reporter cells (Figure 2A).

Bottom Line: We have recently described a novel phenotypic dichotomy within estrogen receptor-positive breast cancer cells; the cell subset responsive to a Sox2 regulatory region (SRR2) reporter (RR cells) are significantly more tumorigenic than the reporter unresponsive (RU) cells.Third, within the CD44(High)/CD24- tumor-initiating cell population derived from MDA-MB-231, RR cells were significantly more tumorigenic than RU cells in an in vivo SCID/Beige xenograft mouse model.To conclude, we described a novel phenotypic heterogeneity within TNBC, and the SRR2 reporter responsiveness is a useful marker for identifying a highly tumorigenic cell subset within the CD44(High)/CD24-tumor-initiating cell population.

View Article: PubMed Central - PubMed

Affiliation: Department of Oncology, University of Alberta, Edmonton, Alberta, Canada.

ABSTRACT
We have recently described a novel phenotypic dichotomy within estrogen receptor-positive breast cancer cells; the cell subset responsive to a Sox2 regulatory region (SRR2) reporter (RR cells) are significantly more tumorigenic than the reporter unresponsive (RU) cells. Here, we report that a similar phenomenon also exists in triple negative breast cancer (TNBC), with RR cells more tumorigenic than RU cells. First, examination of all 3 TNBC cell lines stably infected with the SRR2 reporter revealed the presence of a cell subset exhibiting reporter activity. Second, RU and RR cells purified by flow cytometry showed that RR cells expressed higher levels of CD44, generated more spheres in a limiting dilution mammosphere formation assay, and formed larger and more complex structures in Matrigel. Third, within the CD44(High)/CD24- tumor-initiating cell population derived from MDA-MB-231, RR cells were significantly more tumorigenic than RU cells in an in vivo SCID/Beige xenograft mouse model. Examination of 4 TNBC tumors from patients also revealed the presence of a RR cell subset, ranging from 1.1-3.8%. To conclude, we described a novel phenotypic heterogeneity within TNBC, and the SRR2 reporter responsiveness is a useful marker for identifying a highly tumorigenic cell subset within the CD44(High)/CD24-tumor-initiating cell population.

No MeSH data available.


Related in: MedlinePlus