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Targeted inactivation of β1 integrin induces β3 integrin switching, which drives breast cancer metastasis by TGF-β.

Parvani JG, Galliher-Beckley AJ, Schiemann BJ, Schiemann WP - Mol. Biol. Cell (2013)

Bottom Line: We demonstrate that inactivation of β1 integrin impairs TGF-β from stimulating the motility of normal and malignant mammary epithelial cells (MECs) and elicits robust compensatory expression of β3 integrin solely in malignant MECs, but not in their normal counterparts.Compensatory β3 integrin expression also 1) enhances the growth of malignant MECs in rigid and compliant three-dimensional organotypic cultures and 2) restores the induction of the EMT phenotypes by TGF-β.Of importance, compensatory expression of β3 integrin rescues the growth and pulmonary metastasis of β1 integrin-deficient 4T1 tumors in mice, a process that is prevented by genetic depletion or functional inactivation of β3 integrin.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, Case Western Reserve University, Cleveland, OH 44106 Department of Anatomy and Physiology, College of Veterinary Medicine, Kansas State University, Manhattan, KS 66506 Case Comprehensive Cancer Center, Case Western Reserve University, Cleveland, OH 44106.

ABSTRACT
Mammary tumorigenesis and epithelial-mesenchymal transition (EMT) programs cooperate in converting transforming growth factor-β (TGF-β) from a suppressor to a promoter of breast cancer metastasis. Although previous reports associated β1 and β3 integrins with TGF-β stimulation of EMT and metastasis, the functional interplay and plasticity exhibited by these adhesion molecules in shaping the oncogenic activities of TGF-β remain unknown. We demonstrate that inactivation of β1 integrin impairs TGF-β from stimulating the motility of normal and malignant mammary epithelial cells (MECs) and elicits robust compensatory expression of β3 integrin solely in malignant MECs, but not in their normal counterparts. Compensatory β3 integrin expression also 1) enhances the growth of malignant MECs in rigid and compliant three-dimensional organotypic cultures and 2) restores the induction of the EMT phenotypes by TGF-β. Of importance, compensatory expression of β3 integrin rescues the growth and pulmonary metastasis of β1 integrin-deficient 4T1 tumors in mice, a process that is prevented by genetic depletion or functional inactivation of β3 integrin. Collectively our findings demonstrate that inactivation of β1 integrin elicits metastatic progression via a β3 integrin-specific mechanism, indicating that dual β1 and β3 integrin targeting is necessary to alleviate metastatic disease in breast cancer patients.

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Compensatory β3 integrin expression is essential in enhancing acinar growth of β1 integrin–deficient 4T1 cells. (A, B) Parental (scram) and β1 integrin–deficient 4T1 cells were propagated in rigid 3D-organotypic cultures (3 mg/ml type I collagen) for 4 d, at which point differences in organoid growth and morphology were monitored by phase contrast microscopy (50×; A) and longitudinal bioluminescence (B). Data are mean (±SE) of three independent experiments completed in triplicate. (C, D) Parental (scram) and β1 integrin–deficient 4T1 cells were propagated in compliant 3D-organotypic cultures for 12 d in the absence or presence of the neutralizing αvβ3 integrin antibody LM609 (15 μg/ml). The growth and morphology of the resulting organoids were monitored by phase contrast microscopy (50×; C) and longitudinal bioluminescence (D). Data are mean (±SE) of three independent experiments completed in triplicate (*p < 0.035).
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Figure 6: Compensatory β3 integrin expression is essential in enhancing acinar growth of β1 integrin–deficient 4T1 cells. (A, B) Parental (scram) and β1 integrin–deficient 4T1 cells were propagated in rigid 3D-organotypic cultures (3 mg/ml type I collagen) for 4 d, at which point differences in organoid growth and morphology were monitored by phase contrast microscopy (50×; A) and longitudinal bioluminescence (B). Data are mean (±SE) of three independent experiments completed in triplicate. (C, D) Parental (scram) and β1 integrin–deficient 4T1 cells were propagated in compliant 3D-organotypic cultures for 12 d in the absence or presence of the neutralizing αvβ3 integrin antibody LM609 (15 μg/ml). The growth and morphology of the resulting organoids were monitored by phase contrast microscopy (50×; C) and longitudinal bioluminescence (D). Data are mean (±SE) of three independent experiments completed in triplicate (*p < 0.035).

Mentions: To determine the functional implications of β1 → β3 integrin switching in our mouse models of triple-negative breast cancer, we first propagated parental and β1 integrin–deficient 4T1 cells in rigid, collagen-rich 3D-organotypic cultures to mimic their growth in primary tumor microenvironments (Butcher et al., 2009; Erler and Weaver, 2009; Taylor et al., 2011). In doing so, we observed that parental (scram) 4T1 cells formed highly branched structures, as opposed to those formed by their β1 integrin–deficient counterparts (Figure 6A), which also grew significantly faster than parental cells in these same rigid microenvironments (Figure 6B). Of interest, the growth dynamics of both 4T1 derivatives were identical upon being propagated in compliant 3D-organotypic cultures (Figure 6C). Of importance, administering neutralizing αvβ3 integrin antibodies (LM609) significantly inhibited the growth of β1 integrin–deficient 4T1 cells in compliant 3D-organotypic microenvironments (Figure 6, C and D). Collectively these findings indicate that inactivation of β1 integrin confers triple-negative breast cancers a selective growth advantage in collagen-rich primary tumor microenvironments, as well as in pulmonary microenvironments in part via compensatory β1 → β3 integrin switching.


Targeted inactivation of β1 integrin induces β3 integrin switching, which drives breast cancer metastasis by TGF-β.

Parvani JG, Galliher-Beckley AJ, Schiemann BJ, Schiemann WP - Mol. Biol. Cell (2013)

Compensatory β3 integrin expression is essential in enhancing acinar growth of β1 integrin–deficient 4T1 cells. (A, B) Parental (scram) and β1 integrin–deficient 4T1 cells were propagated in rigid 3D-organotypic cultures (3 mg/ml type I collagen) for 4 d, at which point differences in organoid growth and morphology were monitored by phase contrast microscopy (50×; A) and longitudinal bioluminescence (B). Data are mean (±SE) of three independent experiments completed in triplicate. (C, D) Parental (scram) and β1 integrin–deficient 4T1 cells were propagated in compliant 3D-organotypic cultures for 12 d in the absence or presence of the neutralizing αvβ3 integrin antibody LM609 (15 μg/ml). The growth and morphology of the resulting organoids were monitored by phase contrast microscopy (50×; C) and longitudinal bioluminescence (D). Data are mean (±SE) of three independent experiments completed in triplicate (*p < 0.035).
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Related In: Results  -  Collection

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Figure 6: Compensatory β3 integrin expression is essential in enhancing acinar growth of β1 integrin–deficient 4T1 cells. (A, B) Parental (scram) and β1 integrin–deficient 4T1 cells were propagated in rigid 3D-organotypic cultures (3 mg/ml type I collagen) for 4 d, at which point differences in organoid growth and morphology were monitored by phase contrast microscopy (50×; A) and longitudinal bioluminescence (B). Data are mean (±SE) of three independent experiments completed in triplicate. (C, D) Parental (scram) and β1 integrin–deficient 4T1 cells were propagated in compliant 3D-organotypic cultures for 12 d in the absence or presence of the neutralizing αvβ3 integrin antibody LM609 (15 μg/ml). The growth and morphology of the resulting organoids were monitored by phase contrast microscopy (50×; C) and longitudinal bioluminescence (D). Data are mean (±SE) of three independent experiments completed in triplicate (*p < 0.035).
Mentions: To determine the functional implications of β1 → β3 integrin switching in our mouse models of triple-negative breast cancer, we first propagated parental and β1 integrin–deficient 4T1 cells in rigid, collagen-rich 3D-organotypic cultures to mimic their growth in primary tumor microenvironments (Butcher et al., 2009; Erler and Weaver, 2009; Taylor et al., 2011). In doing so, we observed that parental (scram) 4T1 cells formed highly branched structures, as opposed to those formed by their β1 integrin–deficient counterparts (Figure 6A), which also grew significantly faster than parental cells in these same rigid microenvironments (Figure 6B). Of interest, the growth dynamics of both 4T1 derivatives were identical upon being propagated in compliant 3D-organotypic cultures (Figure 6C). Of importance, administering neutralizing αvβ3 integrin antibodies (LM609) significantly inhibited the growth of β1 integrin–deficient 4T1 cells in compliant 3D-organotypic microenvironments (Figure 6, C and D). Collectively these findings indicate that inactivation of β1 integrin confers triple-negative breast cancers a selective growth advantage in collagen-rich primary tumor microenvironments, as well as in pulmonary microenvironments in part via compensatory β1 → β3 integrin switching.

Bottom Line: We demonstrate that inactivation of β1 integrin impairs TGF-β from stimulating the motility of normal and malignant mammary epithelial cells (MECs) and elicits robust compensatory expression of β3 integrin solely in malignant MECs, but not in their normal counterparts.Compensatory β3 integrin expression also 1) enhances the growth of malignant MECs in rigid and compliant three-dimensional organotypic cultures and 2) restores the induction of the EMT phenotypes by TGF-β.Of importance, compensatory expression of β3 integrin rescues the growth and pulmonary metastasis of β1 integrin-deficient 4T1 tumors in mice, a process that is prevented by genetic depletion or functional inactivation of β3 integrin.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, Case Western Reserve University, Cleveland, OH 44106 Department of Anatomy and Physiology, College of Veterinary Medicine, Kansas State University, Manhattan, KS 66506 Case Comprehensive Cancer Center, Case Western Reserve University, Cleveland, OH 44106.

ABSTRACT
Mammary tumorigenesis and epithelial-mesenchymal transition (EMT) programs cooperate in converting transforming growth factor-β (TGF-β) from a suppressor to a promoter of breast cancer metastasis. Although previous reports associated β1 and β3 integrins with TGF-β stimulation of EMT and metastasis, the functional interplay and plasticity exhibited by these adhesion molecules in shaping the oncogenic activities of TGF-β remain unknown. We demonstrate that inactivation of β1 integrin impairs TGF-β from stimulating the motility of normal and malignant mammary epithelial cells (MECs) and elicits robust compensatory expression of β3 integrin solely in malignant MECs, but not in their normal counterparts. Compensatory β3 integrin expression also 1) enhances the growth of malignant MECs in rigid and compliant three-dimensional organotypic cultures and 2) restores the induction of the EMT phenotypes by TGF-β. Of importance, compensatory expression of β3 integrin rescues the growth and pulmonary metastasis of β1 integrin-deficient 4T1 tumors in mice, a process that is prevented by genetic depletion or functional inactivation of β3 integrin. Collectively our findings demonstrate that inactivation of β1 integrin elicits metastatic progression via a β3 integrin-specific mechanism, indicating that dual β1 and β3 integrin targeting is necessary to alleviate metastatic disease in breast cancer patients.

Show MeSH
Related in: MedlinePlus