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mTORC1 maintains the tumorigenicity of SSEA-4(+) high-grade osteosarcoma.

Zhang W, Ding ML, Zhang JN, Qiu JR, Shen YH, Ding XY, Deng LF, Zhang WB, Zhu J - Sci Rep (2015)

Bottom Line: Here we show that in certain high-grade osteosarcomas, immature SSEA-4(+) tumor cells represent a subset of tumor-initiating cells (TICs) whose pool size is maintained by mTORC1 activity. mTORC1 supports not only SSEA-4(+) cell self-renewal through S6K but also the regeneration of SSEA-4(+) TICs by SSEA-4(-) osteosarcoma cell dedifferentiation.Mechanistically, active mTORC1 is required to prevent a likely upregulation of the cell-cycle inhibitor p27 independently of p53 or Rb activation, which otherwise effectively drives the terminal differentiation of SSEA-4(-) osteosarcoma cells at the expense of dedifferentiation.Thus, mTORC1 is shown to critically regulate the retention of tumorigenicity versus differentiation in discrete differentiation phases in SSEA-4(+) TICs and their progeny.

View Article: PubMed Central - PubMed

Affiliation: 1] State Key Laboratory for Medical Genomics, Shanghai Institute of Hematology and Collaborative Innovation Center of Hematology, Rui-Jin Hospital affiliated to Shanghai Jiao-Tong University School of Medicine, Shanghai 200025, People's Republic of China [2] Collaborative Innovation Center of Systems Biomedicine, Shanghai 200025, People's Republic of China.

ABSTRACT
Inactivation of p53 and/or Rb pathways restrains osteoblasts from cell-cycle exit and terminal differentiation, which underpins osteosarcoma formation coupled with dedifferentiation. Recently, the level of p-S6K was shown to independently predict the prognosis for osteosarcomas, while the reason behind this is not understood. Here we show that in certain high-grade osteosarcomas, immature SSEA-4(+) tumor cells represent a subset of tumor-initiating cells (TICs) whose pool size is maintained by mTORC1 activity. mTORC1 supports not only SSEA-4(+) cell self-renewal through S6K but also the regeneration of SSEA-4(+) TICs by SSEA-4(-) osteosarcoma cell dedifferentiation. Mechanistically, active mTORC1 is required to prevent a likely upregulation of the cell-cycle inhibitor p27 independently of p53 or Rb activation, which otherwise effectively drives the terminal differentiation of SSEA-4(-) osteosarcoma cells at the expense of dedifferentiation. Thus, mTORC1 is shown to critically regulate the retention of tumorigenicity versus differentiation in discrete differentiation phases in SSEA-4(+) TICs and their progeny.

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SSEA-4+ TICs are Responsible for the Clinical Progression of a Distinct Subtype of High-grade Osteosarcomas.(a) Overall survival of patients with discrete SSEA-4 staining intensities in osteosarcoma cases, as measured before (left panel) or after (right panel) the first round of chemotherapy. P values: compared with the SSEA-4− subgroup; n: patient number. (b) Oct3/4 expression assayed on primary specimens of osteosarcoma that contained or did not contain SSEA-4+ TICs. Percentages of Oct3/4+ cells are shown in the upper-right corner. Six representative samples are shown. (c) Cryopreserved sections of SSEA-4+ xenografts were co-stained with fluorescent antibodies against Oct3/4 or SSEA-4. Three representative samples are shown. Red arrows indicate SSEA-4+ cells. (d) Upper panel, SSEA-4 staining before and after the first round of chemotherapy in one representative case. Bottom panel, SSEA-4-staining intensities of osteosarcoma samples from 19 patients measured before and after chemotherapy. Samples from the same patient are paired with lines. (e) Relative distribution rates of SSEA-4−, SSEA-41+, SSEA-42+, and SSEA-43+ osteosarcoma samples among those obtained from primary or metastatic sites.
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f2: SSEA-4+ TICs are Responsible for the Clinical Progression of a Distinct Subtype of High-grade Osteosarcomas.(a) Overall survival of patients with discrete SSEA-4 staining intensities in osteosarcoma cases, as measured before (left panel) or after (right panel) the first round of chemotherapy. P values: compared with the SSEA-4− subgroup; n: patient number. (b) Oct3/4 expression assayed on primary specimens of osteosarcoma that contained or did not contain SSEA-4+ TICs. Percentages of Oct3/4+ cells are shown in the upper-right corner. Six representative samples are shown. (c) Cryopreserved sections of SSEA-4+ xenografts were co-stained with fluorescent antibodies against Oct3/4 or SSEA-4. Three representative samples are shown. Red arrows indicate SSEA-4+ cells. (d) Upper panel, SSEA-4 staining before and after the first round of chemotherapy in one representative case. Bottom panel, SSEA-4-staining intensities of osteosarcoma samples from 19 patients measured before and after chemotherapy. Samples from the same patient are paired with lines. (e) Relative distribution rates of SSEA-4−, SSEA-41+, SSEA-42+, and SSEA-43+ osteosarcoma samples among those obtained from primary or metastatic sites.

Mentions: SSEA-4+ osteosarcoma cells were readily detectable only in a small fraction (8/21) of primary osteosarcoma specimens (Fig. 1c, d), which prompted us to test whether the osteosarcoma cases containing SSEA-4+ TICs represent a subtype of osteosarcoma distinct from the majority of SSEA-4neg cases. To address this, we performed a retrospective analysis of a cohort of osteosarcoma cases collected over >10 years. Remarkably, the frequency of SSEA-4+ TICs alone, as indicated by immunohistochemical staining (arbitrarily determined as -, 1+, 2+, or 3+; see Supplementary Fig. 2a) before or after the first round of chemotherapy, correlated inversely with the overall-survival probabilities of patients (Fig. 2a). Strikingly, >90% of SSEA-4neg cases but no SSEA-42+–3+ cases survived more than 10 years after diagnosis. Notably, transcription factor Oct3/4, the expression of which marks MSCs and was reported in certain cultivated osteosarcoma cells to associate with a gain of tumorigenicity and probably reprogramming events283031, was consistently and abundantly expressed in bulk tissue cells of SSEA-4+2–3 primary specimens (n = 26) wherein SSEA-4+ TICs constituted a small subfraction of Oct3/4+ osteosarcoma cells (Fig. 2b, c). In striking contrast, no Oct3/4 expression was detected in eight SSEA-4neg cases examined (Fig. 2b, Supplementary Fig. 2b, c), thus indicating that SSEA-4+ TICs but not TICs from other subtypes of osteosarcoma have been reprogrammed to a highly immature status comparable to MSCs or even ES cells in certain molecular programs32.


mTORC1 maintains the tumorigenicity of SSEA-4(+) high-grade osteosarcoma.

Zhang W, Ding ML, Zhang JN, Qiu JR, Shen YH, Ding XY, Deng LF, Zhang WB, Zhu J - Sci Rep (2015)

SSEA-4+ TICs are Responsible for the Clinical Progression of a Distinct Subtype of High-grade Osteosarcomas.(a) Overall survival of patients with discrete SSEA-4 staining intensities in osteosarcoma cases, as measured before (left panel) or after (right panel) the first round of chemotherapy. P values: compared with the SSEA-4− subgroup; n: patient number. (b) Oct3/4 expression assayed on primary specimens of osteosarcoma that contained or did not contain SSEA-4+ TICs. Percentages of Oct3/4+ cells are shown in the upper-right corner. Six representative samples are shown. (c) Cryopreserved sections of SSEA-4+ xenografts were co-stained with fluorescent antibodies against Oct3/4 or SSEA-4. Three representative samples are shown. Red arrows indicate SSEA-4+ cells. (d) Upper panel, SSEA-4 staining before and after the first round of chemotherapy in one representative case. Bottom panel, SSEA-4-staining intensities of osteosarcoma samples from 19 patients measured before and after chemotherapy. Samples from the same patient are paired with lines. (e) Relative distribution rates of SSEA-4−, SSEA-41+, SSEA-42+, and SSEA-43+ osteosarcoma samples among those obtained from primary or metastatic sites.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4389812&req=5

f2: SSEA-4+ TICs are Responsible for the Clinical Progression of a Distinct Subtype of High-grade Osteosarcomas.(a) Overall survival of patients with discrete SSEA-4 staining intensities in osteosarcoma cases, as measured before (left panel) or after (right panel) the first round of chemotherapy. P values: compared with the SSEA-4− subgroup; n: patient number. (b) Oct3/4 expression assayed on primary specimens of osteosarcoma that contained or did not contain SSEA-4+ TICs. Percentages of Oct3/4+ cells are shown in the upper-right corner. Six representative samples are shown. (c) Cryopreserved sections of SSEA-4+ xenografts were co-stained with fluorescent antibodies against Oct3/4 or SSEA-4. Three representative samples are shown. Red arrows indicate SSEA-4+ cells. (d) Upper panel, SSEA-4 staining before and after the first round of chemotherapy in one representative case. Bottom panel, SSEA-4-staining intensities of osteosarcoma samples from 19 patients measured before and after chemotherapy. Samples from the same patient are paired with lines. (e) Relative distribution rates of SSEA-4−, SSEA-41+, SSEA-42+, and SSEA-43+ osteosarcoma samples among those obtained from primary or metastatic sites.
Mentions: SSEA-4+ osteosarcoma cells were readily detectable only in a small fraction (8/21) of primary osteosarcoma specimens (Fig. 1c, d), which prompted us to test whether the osteosarcoma cases containing SSEA-4+ TICs represent a subtype of osteosarcoma distinct from the majority of SSEA-4neg cases. To address this, we performed a retrospective analysis of a cohort of osteosarcoma cases collected over >10 years. Remarkably, the frequency of SSEA-4+ TICs alone, as indicated by immunohistochemical staining (arbitrarily determined as -, 1+, 2+, or 3+; see Supplementary Fig. 2a) before or after the first round of chemotherapy, correlated inversely with the overall-survival probabilities of patients (Fig. 2a). Strikingly, >90% of SSEA-4neg cases but no SSEA-42+–3+ cases survived more than 10 years after diagnosis. Notably, transcription factor Oct3/4, the expression of which marks MSCs and was reported in certain cultivated osteosarcoma cells to associate with a gain of tumorigenicity and probably reprogramming events283031, was consistently and abundantly expressed in bulk tissue cells of SSEA-4+2–3 primary specimens (n = 26) wherein SSEA-4+ TICs constituted a small subfraction of Oct3/4+ osteosarcoma cells (Fig. 2b, c). In striking contrast, no Oct3/4 expression was detected in eight SSEA-4neg cases examined (Fig. 2b, Supplementary Fig. 2b, c), thus indicating that SSEA-4+ TICs but not TICs from other subtypes of osteosarcoma have been reprogrammed to a highly immature status comparable to MSCs or even ES cells in certain molecular programs32.

Bottom Line: Here we show that in certain high-grade osteosarcomas, immature SSEA-4(+) tumor cells represent a subset of tumor-initiating cells (TICs) whose pool size is maintained by mTORC1 activity. mTORC1 supports not only SSEA-4(+) cell self-renewal through S6K but also the regeneration of SSEA-4(+) TICs by SSEA-4(-) osteosarcoma cell dedifferentiation.Mechanistically, active mTORC1 is required to prevent a likely upregulation of the cell-cycle inhibitor p27 independently of p53 or Rb activation, which otherwise effectively drives the terminal differentiation of SSEA-4(-) osteosarcoma cells at the expense of dedifferentiation.Thus, mTORC1 is shown to critically regulate the retention of tumorigenicity versus differentiation in discrete differentiation phases in SSEA-4(+) TICs and their progeny.

View Article: PubMed Central - PubMed

Affiliation: 1] State Key Laboratory for Medical Genomics, Shanghai Institute of Hematology and Collaborative Innovation Center of Hematology, Rui-Jin Hospital affiliated to Shanghai Jiao-Tong University School of Medicine, Shanghai 200025, People's Republic of China [2] Collaborative Innovation Center of Systems Biomedicine, Shanghai 200025, People's Republic of China.

ABSTRACT
Inactivation of p53 and/or Rb pathways restrains osteoblasts from cell-cycle exit and terminal differentiation, which underpins osteosarcoma formation coupled with dedifferentiation. Recently, the level of p-S6K was shown to independently predict the prognosis for osteosarcomas, while the reason behind this is not understood. Here we show that in certain high-grade osteosarcomas, immature SSEA-4(+) tumor cells represent a subset of tumor-initiating cells (TICs) whose pool size is maintained by mTORC1 activity. mTORC1 supports not only SSEA-4(+) cell self-renewal through S6K but also the regeneration of SSEA-4(+) TICs by SSEA-4(-) osteosarcoma cell dedifferentiation. Mechanistically, active mTORC1 is required to prevent a likely upregulation of the cell-cycle inhibitor p27 independently of p53 or Rb activation, which otherwise effectively drives the terminal differentiation of SSEA-4(-) osteosarcoma cells at the expense of dedifferentiation. Thus, mTORC1 is shown to critically regulate the retention of tumorigenicity versus differentiation in discrete differentiation phases in SSEA-4(+) TICs and their progeny.

Show MeSH
Related in: MedlinePlus