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ELF5 Drives Lung Metastasis in Luminal Breast Cancer through Recruitment of Gr1+ CD11b+ Myeloid-Derived Suppressor Cells.

Gallego-Ortega D, Ledger A, Roden DL, Law AM, Magenau A, Kikhtyak Z, Cho C, Allerdice SL, Lee HJ, Valdes-Mora F, Herrmann D, Salomon R, Young AI, Lee BY, Sergio CM, Kaplan W, Piggin C, Conway JR, Rabinovich B, Millar EK, Oakes SR, Chtanova T, Swarbrick A, Naylor MJ, O'Toole S, Green AR, Timpson P, Gee JM, Ellis IO, Clark SJ, Ormandy CJ - PLoS Biol. (2015)

Bottom Line: Myeloid-derived suppressor cells, a group of immature neutrophils recently identified as mediators of vasculogenesis and metastasis, were recruited to the tumor in response to ELF5.Depletion of these cells using specific Ly6G antibodies prevented ELF5 from driving vasculogenesis and metastasis.Thus, in the MMTV-PyMT mouse mammary model, increased ELF5 levels drive metastasis by co-opting the innate immune system.

View Article: PubMed Central - PubMed

Affiliation: Garvan Institute of Medical Research and The Kinghorn Cancer Centre, Darlinghurst, New South Wales, Australia.

ABSTRACT
During pregnancy, the ETS transcription factor ELF5 establishes the milk-secreting alveolar cell lineage by driving a cell fate decision of the mammary luminal progenitor cell. In breast cancer, ELF5 is a key transcriptional determinant of tumor subtype and has been implicated in the development of insensitivity to anti-estrogen therapy. In the mouse mammary tumor virus-Polyoma Middle T (MMTV-PyMT) model of luminal breast cancer, induction of ELF5 levels increased leukocyte infiltration, angiogenesis, and blood vessel permeability in primary tumors and greatly increased the size and number of lung metastasis. Myeloid-derived suppressor cells, a group of immature neutrophils recently identified as mediators of vasculogenesis and metastasis, were recruited to the tumor in response to ELF5. Depletion of these cells using specific Ly6G antibodies prevented ELF5 from driving vasculogenesis and metastasis. Expression signatures in luminal A breast cancers indicated that increased myeloid cell invasion and inflammation were correlated with ELF5 expression, and increased ELF5 immunohistochemical staining predicted much shorter metastasis-free and overall survival of luminal A patients, defining a group who experienced unexpectedly early disease progression. Thus, in the MMTV-PyMT mouse mammary model, increased ELF5 levels drive metastasis by co-opting the innate immune system. As ELF5 has been previously implicated in the development of antiestrogen resistance, this finding implicates ELF5 as a defining factor in the acquisition of the key aspects of the lethal phenotype in luminal A breast cancer.

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ELF5 expression in the tumor epithelium increases metastases to the lungs.Panel A, appearance of lungs from a control PyMT animal following long term (8 wk) DOX treatment. Panel B, H&E histology of lungs in Panel A. Panels C and D, examples of H&E histology of lungs from control PyMT mice receiving short term (2 wk) DOX treatment. Panel E, appearance of lungs following long term ELF5 expression. Panel F, H&E histology of the lungs in Panel E. Panel G and H, H&E histology of lungs from mice receiving short term induction of ELF5. Panel I, visualization of EGFP of the lungs in Panel E. Panel J, example of IHC staining for ELF5 in a PyMT/ELF5 lung metastasis. Panel K, relationship between the size of an individual lung lesion and the IHC score for ELF5 level (combining intensity and percent positivity). Panel L and M, quantification of the number of metastases in the lungs of the mice with the indicated genotypes after long or short term DOX exposure respectively. Panel N, metastatic behavior of the indicated genotypes expressed as an area. Panel O, PyMT expression measured by qPCR in the blood of mice of the indicated genotypes. Panel P, comparison to the number of metastases driven by pregnancy. Labels are pregnancy (preg.) and iparous (ip.). Raw data for panels K, L, M, N, O, and P can be found at S3 Data.
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pbio.1002330.g004: ELF5 expression in the tumor epithelium increases metastases to the lungs.Panel A, appearance of lungs from a control PyMT animal following long term (8 wk) DOX treatment. Panel B, H&E histology of lungs in Panel A. Panels C and D, examples of H&E histology of lungs from control PyMT mice receiving short term (2 wk) DOX treatment. Panel E, appearance of lungs following long term ELF5 expression. Panel F, H&E histology of the lungs in Panel E. Panel G and H, H&E histology of lungs from mice receiving short term induction of ELF5. Panel I, visualization of EGFP of the lungs in Panel E. Panel J, example of IHC staining for ELF5 in a PyMT/ELF5 lung metastasis. Panel K, relationship between the size of an individual lung lesion and the IHC score for ELF5 level (combining intensity and percent positivity). Panel L and M, quantification of the number of metastases in the lungs of the mice with the indicated genotypes after long or short term DOX exposure respectively. Panel N, metastatic behavior of the indicated genotypes expressed as an area. Panel O, PyMT expression measured by qPCR in the blood of mice of the indicated genotypes. Panel P, comparison to the number of metastases driven by pregnancy. Labels are pregnancy (preg.) and iparous (ip.). Raw data for panels K, L, M, N, O, and P can be found at S3 Data.

Mentions: We examined the effect of the induction of ELF5 on the metastatic behavior of the PyMT model. In control animals, constitutive PyMT expression produced no visible lung metastatic nodules by the time the primary tumors reached the ethical endpoint of 10% body weight (Fig 4A), but small metastases within the lungs were detectable by H&E histology (Fig 4B). DOX administration in control animals had no effect on metastasis (Fig 4C and 4D). Induction of ELF5 from 6 wk of age resulted in a dramatic increase in metastasis to the lungs, now visible as numerous nodules on the surface of the lung at the ethical endpoint (Fig 4E) and large and numerous metastases within the lungs by H&E histology (Fig 4F). Induction of ELF5 for 2 wk once tumors were palpable also increased the size and number of detectable lung metastases (Fig 4G and 4H) but with more variable penetrance between animals compared with longer DOX treatment. Most of these metastases expressed ELF5, observed by visualization of EGFP (Fig 4I) and by ELF5 IHC (Fig 4J). Quantification showed a positive correlation between the size of the metastatic lesion and the level of ELF5 protein (Fig 4K). Unlike the primary tumors the metastases showed no regions of hemorrhage. Quantification of H&E stained sections showed statistically significant increases in the number of lung metastases (Fig 4L and 4M). Measurement of metastatic area produced similar results (Fig 4N). Induction of ELF5 greatly increased the amount of PyMT-mRNA present in blood (Fig 4O), suggesting increased numbers of circulating tumor cells. Elf5 is a master regulator of the development and remodeling of the mammary epithelium during pregnancy. During this period Elf5 is intensively expressed. We found that the metastasis-promoting effect of Elf5 was comparable to that produced by pregnancy in this model (Fig 4P).


ELF5 Drives Lung Metastasis in Luminal Breast Cancer through Recruitment of Gr1+ CD11b+ Myeloid-Derived Suppressor Cells.

Gallego-Ortega D, Ledger A, Roden DL, Law AM, Magenau A, Kikhtyak Z, Cho C, Allerdice SL, Lee HJ, Valdes-Mora F, Herrmann D, Salomon R, Young AI, Lee BY, Sergio CM, Kaplan W, Piggin C, Conway JR, Rabinovich B, Millar EK, Oakes SR, Chtanova T, Swarbrick A, Naylor MJ, O'Toole S, Green AR, Timpson P, Gee JM, Ellis IO, Clark SJ, Ormandy CJ - PLoS Biol. (2015)

ELF5 expression in the tumor epithelium increases metastases to the lungs.Panel A, appearance of lungs from a control PyMT animal following long term (8 wk) DOX treatment. Panel B, H&E histology of lungs in Panel A. Panels C and D, examples of H&E histology of lungs from control PyMT mice receiving short term (2 wk) DOX treatment. Panel E, appearance of lungs following long term ELF5 expression. Panel F, H&E histology of the lungs in Panel E. Panel G and H, H&E histology of lungs from mice receiving short term induction of ELF5. Panel I, visualization of EGFP of the lungs in Panel E. Panel J, example of IHC staining for ELF5 in a PyMT/ELF5 lung metastasis. Panel K, relationship between the size of an individual lung lesion and the IHC score for ELF5 level (combining intensity and percent positivity). Panel L and M, quantification of the number of metastases in the lungs of the mice with the indicated genotypes after long or short term DOX exposure respectively. Panel N, metastatic behavior of the indicated genotypes expressed as an area. Panel O, PyMT expression measured by qPCR in the blood of mice of the indicated genotypes. Panel P, comparison to the number of metastases driven by pregnancy. Labels are pregnancy (preg.) and iparous (ip.). Raw data for panels K, L, M, N, O, and P can be found at S3 Data.
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pbio.1002330.g004: ELF5 expression in the tumor epithelium increases metastases to the lungs.Panel A, appearance of lungs from a control PyMT animal following long term (8 wk) DOX treatment. Panel B, H&E histology of lungs in Panel A. Panels C and D, examples of H&E histology of lungs from control PyMT mice receiving short term (2 wk) DOX treatment. Panel E, appearance of lungs following long term ELF5 expression. Panel F, H&E histology of the lungs in Panel E. Panel G and H, H&E histology of lungs from mice receiving short term induction of ELF5. Panel I, visualization of EGFP of the lungs in Panel E. Panel J, example of IHC staining for ELF5 in a PyMT/ELF5 lung metastasis. Panel K, relationship between the size of an individual lung lesion and the IHC score for ELF5 level (combining intensity and percent positivity). Panel L and M, quantification of the number of metastases in the lungs of the mice with the indicated genotypes after long or short term DOX exposure respectively. Panel N, metastatic behavior of the indicated genotypes expressed as an area. Panel O, PyMT expression measured by qPCR in the blood of mice of the indicated genotypes. Panel P, comparison to the number of metastases driven by pregnancy. Labels are pregnancy (preg.) and iparous (ip.). Raw data for panels K, L, M, N, O, and P can be found at S3 Data.
Mentions: We examined the effect of the induction of ELF5 on the metastatic behavior of the PyMT model. In control animals, constitutive PyMT expression produced no visible lung metastatic nodules by the time the primary tumors reached the ethical endpoint of 10% body weight (Fig 4A), but small metastases within the lungs were detectable by H&E histology (Fig 4B). DOX administration in control animals had no effect on metastasis (Fig 4C and 4D). Induction of ELF5 from 6 wk of age resulted in a dramatic increase in metastasis to the lungs, now visible as numerous nodules on the surface of the lung at the ethical endpoint (Fig 4E) and large and numerous metastases within the lungs by H&E histology (Fig 4F). Induction of ELF5 for 2 wk once tumors were palpable also increased the size and number of detectable lung metastases (Fig 4G and 4H) but with more variable penetrance between animals compared with longer DOX treatment. Most of these metastases expressed ELF5, observed by visualization of EGFP (Fig 4I) and by ELF5 IHC (Fig 4J). Quantification showed a positive correlation between the size of the metastatic lesion and the level of ELF5 protein (Fig 4K). Unlike the primary tumors the metastases showed no regions of hemorrhage. Quantification of H&E stained sections showed statistically significant increases in the number of lung metastases (Fig 4L and 4M). Measurement of metastatic area produced similar results (Fig 4N). Induction of ELF5 greatly increased the amount of PyMT-mRNA present in blood (Fig 4O), suggesting increased numbers of circulating tumor cells. Elf5 is a master regulator of the development and remodeling of the mammary epithelium during pregnancy. During this period Elf5 is intensively expressed. We found that the metastasis-promoting effect of Elf5 was comparable to that produced by pregnancy in this model (Fig 4P).

Bottom Line: Myeloid-derived suppressor cells, a group of immature neutrophils recently identified as mediators of vasculogenesis and metastasis, were recruited to the tumor in response to ELF5.Depletion of these cells using specific Ly6G antibodies prevented ELF5 from driving vasculogenesis and metastasis.Thus, in the MMTV-PyMT mouse mammary model, increased ELF5 levels drive metastasis by co-opting the innate immune system.

View Article: PubMed Central - PubMed

Affiliation: Garvan Institute of Medical Research and The Kinghorn Cancer Centre, Darlinghurst, New South Wales, Australia.

ABSTRACT
During pregnancy, the ETS transcription factor ELF5 establishes the milk-secreting alveolar cell lineage by driving a cell fate decision of the mammary luminal progenitor cell. In breast cancer, ELF5 is a key transcriptional determinant of tumor subtype and has been implicated in the development of insensitivity to anti-estrogen therapy. In the mouse mammary tumor virus-Polyoma Middle T (MMTV-PyMT) model of luminal breast cancer, induction of ELF5 levels increased leukocyte infiltration, angiogenesis, and blood vessel permeability in primary tumors and greatly increased the size and number of lung metastasis. Myeloid-derived suppressor cells, a group of immature neutrophils recently identified as mediators of vasculogenesis and metastasis, were recruited to the tumor in response to ELF5. Depletion of these cells using specific Ly6G antibodies prevented ELF5 from driving vasculogenesis and metastasis. Expression signatures in luminal A breast cancers indicated that increased myeloid cell invasion and inflammation were correlated with ELF5 expression, and increased ELF5 immunohistochemical staining predicted much shorter metastasis-free and overall survival of luminal A patients, defining a group who experienced unexpectedly early disease progression. Thus, in the MMTV-PyMT mouse mammary model, increased ELF5 levels drive metastasis by co-opting the innate immune system. As ELF5 has been previously implicated in the development of antiestrogen resistance, this finding implicates ELF5 as a defining factor in the acquisition of the key aspects of the lethal phenotype in luminal A breast cancer.

Show MeSH
Related in: MedlinePlus