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Adipose cells promote resistance of breast cancer cells to trastuzumab-mediated antibody-dependent cellular cytotoxicity.

Duong MN, Cleret A, Matera EL, Chettab K, Mathé D, Valsesia-Wittmann S, Clémenceau B, Dumontet C - Breast Cancer Res. (2015)

Bottom Line: The results were validated in vivo in a mouse xenograft model.Using a transcriptomic approach, we found that cancer cells undergo major modifications when exposed to adipocyte-conditioned medium.Collectively, our findings underline the importance of adipose tissue in the resistance to trastuzumab and suggest that approaches targeting the adipocyte-cancer cell crosstalk may help sensitize cancer cells to trastuzumab-based therapy.

View Article: PubMed Central - PubMed

Affiliation: Centre de Recherche en Cancérologie de Lyon (CRCL), INSERM UMR 1052, CNRS 5286, 8 Avenue Rockefeller, 69008, Lyon, France. mn.duong@hotmail.com.

ABSTRACT

Introduction: Trastuzumab has been used in the treatment of human epidermal growth factor receptor 2 (HER2)-expressing breast cancer, but its efficacy is limited by de novo or acquired resistance. Although many mechanisms have been proposed to explain resistance to trastuzumab, little is known concerning the role of the tumor microenvironment. Given the importance of antibody-dependent cellular cytotoxicity (ADCC) in the antitumor effect of trastuzumab and the abundance of adipose tissue in the breast, we investigated the impact of adipocytes on ADCC.

Methods: We set up a coculture system to study the effect of adipocytes on ADCC in vitro. The results were validated in vivo in a mouse xenograft model.

Results: We found that adipocytes, as well as preadipocytes, inhibited trastuzumab-mediated ADCC in HER2-expressing breast cancer cells via the secretion of soluble factors. The inhibition of ADCC was not due to titration or degradation of the antibody. We found that adipose cells decreased the secretion of interferon-γ by natural killer cells, but did not alter natural killer cells' cytotoxicity. Preincubation of breast cancer cells with the conditioned medium derived from adipocytes reduced the sensitivity of cancer cells to ADCC. Using a transcriptomic approach, we found that cancer cells undergo major modifications when exposed to adipocyte-conditioned medium. Importantly, breast tumors grafted next to lipomas displayed resistance to trastuzumab in mouse xenograft models.

Conclusions: Collectively, our findings underline the importance of adipose tissue in the resistance to trastuzumab and suggest that approaches targeting the adipocyte-cancer cell crosstalk may help sensitize cancer cells to trastuzumab-based therapy.

No MeSH data available.


Related in: MedlinePlus

Conditioned media of differentiated human multipotent adipose-derived stem cells increases the resistance of BT-474 cells against ADCC. Human epidermal growth factor receptor 2 (HER2) expression (A) or localization (B) on BT-474 cells after 4-hour incubation with conditioned media of differentiated human multipotent adipose-derived stem cells (#hMADS-CM) or undifferentiated human multipotent adipose-derived stem cells (hMADS-CM) or their control media. Scale bars indicate 10 μm. (C) Antibody-dependent cellular cytotoxicity (ADCC) assays of BT-474 cells preincubated overnight with #hMADS-CM, hMADS-CM or their control media. (D) Kinetic induction of Akt phosphorylation (Phospho Akt) in BT-474 cells exposed to #hMADS-CM or the control medium (#medium) for the indicated times. Quantification of the intensity of the bands is shown. Mean ± SD values of three independent experiments are shown in (A) and (C). Results representative of three independent experiments are shown in (B) and (D). (E) Reversion of #hMADS-induced inhibition of ADCC by temsirolimus. Temsirolimus was added at the beginning of ADCC assays at the indicated concentrations. (F) Genes involved in cell survival in BT-474 cells after exposure to #hMADS-CM. Numbers correspond to fold changes. Vertical rectangle, G protein–coupled receptor; dashed square, growth factor; inverted triangle, kinase; horizontal rectangle, ligand-dependent nuclear receptor; triangle, phosphatase; oval, transcription regulator; trapezoid, transporter; circle, other. Genes in red or green correspond to upregulated or downregulated, respectively. Red lines predict activation; yellow lines indicate inconsistent downstream effect; and gray lines correspond to unpredicted effect. *P < 0.05; **P < 0.01; ns, Not significant.
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Fig5: Conditioned media of differentiated human multipotent adipose-derived stem cells increases the resistance of BT-474 cells against ADCC. Human epidermal growth factor receptor 2 (HER2) expression (A) or localization (B) on BT-474 cells after 4-hour incubation with conditioned media of differentiated human multipotent adipose-derived stem cells (#hMADS-CM) or undifferentiated human multipotent adipose-derived stem cells (hMADS-CM) or their control media. Scale bars indicate 10 μm. (C) Antibody-dependent cellular cytotoxicity (ADCC) assays of BT-474 cells preincubated overnight with #hMADS-CM, hMADS-CM or their control media. (D) Kinetic induction of Akt phosphorylation (Phospho Akt) in BT-474 cells exposed to #hMADS-CM or the control medium (#medium) for the indicated times. Quantification of the intensity of the bands is shown. Mean ± SD values of three independent experiments are shown in (A) and (C). Results representative of three independent experiments are shown in (B) and (D). (E) Reversion of #hMADS-induced inhibition of ADCC by temsirolimus. Temsirolimus was added at the beginning of ADCC assays at the indicated concentrations. (F) Genes involved in cell survival in BT-474 cells after exposure to #hMADS-CM. Numbers correspond to fold changes. Vertical rectangle, G protein–coupled receptor; dashed square, growth factor; inverted triangle, kinase; horizontal rectangle, ligand-dependent nuclear receptor; triangle, phosphatase; oval, transcription regulator; trapezoid, transporter; circle, other. Genes in red or green correspond to upregulated or downregulated, respectively. Red lines predict activation; yellow lines indicate inconsistent downstream effect; and gray lines correspond to unpredicted effect. *P < 0.05; **P < 0.01; ns, Not significant.

Mentions: BT-474 cells were exposed to either #hMADS-CM or control medium for the Figure 5D. Proteins were extracted in radioimmunoprecipitation assay buffer and subjected to SDS-PAGE and immunoblot analysis. The antibodies used were anti-phospho-Akt and anti-Akt (Cell Signaling Technology, Beverly, MA, USA) and anti-tubulin (Sigma-Aldrich).


Adipose cells promote resistance of breast cancer cells to trastuzumab-mediated antibody-dependent cellular cytotoxicity.

Duong MN, Cleret A, Matera EL, Chettab K, Mathé D, Valsesia-Wittmann S, Clémenceau B, Dumontet C - Breast Cancer Res. (2015)

Conditioned media of differentiated human multipotent adipose-derived stem cells increases the resistance of BT-474 cells against ADCC. Human epidermal growth factor receptor 2 (HER2) expression (A) or localization (B) on BT-474 cells after 4-hour incubation with conditioned media of differentiated human multipotent adipose-derived stem cells (#hMADS-CM) or undifferentiated human multipotent adipose-derived stem cells (hMADS-CM) or their control media. Scale bars indicate 10 μm. (C) Antibody-dependent cellular cytotoxicity (ADCC) assays of BT-474 cells preincubated overnight with #hMADS-CM, hMADS-CM or their control media. (D) Kinetic induction of Akt phosphorylation (Phospho Akt) in BT-474 cells exposed to #hMADS-CM or the control medium (#medium) for the indicated times. Quantification of the intensity of the bands is shown. Mean ± SD values of three independent experiments are shown in (A) and (C). Results representative of three independent experiments are shown in (B) and (D). (E) Reversion of #hMADS-induced inhibition of ADCC by temsirolimus. Temsirolimus was added at the beginning of ADCC assays at the indicated concentrations. (F) Genes involved in cell survival in BT-474 cells after exposure to #hMADS-CM. Numbers correspond to fold changes. Vertical rectangle, G protein–coupled receptor; dashed square, growth factor; inverted triangle, kinase; horizontal rectangle, ligand-dependent nuclear receptor; triangle, phosphatase; oval, transcription regulator; trapezoid, transporter; circle, other. Genes in red or green correspond to upregulated or downregulated, respectively. Red lines predict activation; yellow lines indicate inconsistent downstream effect; and gray lines correspond to unpredicted effect. *P < 0.05; **P < 0.01; ns, Not significant.
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Fig5: Conditioned media of differentiated human multipotent adipose-derived stem cells increases the resistance of BT-474 cells against ADCC. Human epidermal growth factor receptor 2 (HER2) expression (A) or localization (B) on BT-474 cells after 4-hour incubation with conditioned media of differentiated human multipotent adipose-derived stem cells (#hMADS-CM) or undifferentiated human multipotent adipose-derived stem cells (hMADS-CM) or their control media. Scale bars indicate 10 μm. (C) Antibody-dependent cellular cytotoxicity (ADCC) assays of BT-474 cells preincubated overnight with #hMADS-CM, hMADS-CM or their control media. (D) Kinetic induction of Akt phosphorylation (Phospho Akt) in BT-474 cells exposed to #hMADS-CM or the control medium (#medium) for the indicated times. Quantification of the intensity of the bands is shown. Mean ± SD values of three independent experiments are shown in (A) and (C). Results representative of three independent experiments are shown in (B) and (D). (E) Reversion of #hMADS-induced inhibition of ADCC by temsirolimus. Temsirolimus was added at the beginning of ADCC assays at the indicated concentrations. (F) Genes involved in cell survival in BT-474 cells after exposure to #hMADS-CM. Numbers correspond to fold changes. Vertical rectangle, G protein–coupled receptor; dashed square, growth factor; inverted triangle, kinase; horizontal rectangle, ligand-dependent nuclear receptor; triangle, phosphatase; oval, transcription regulator; trapezoid, transporter; circle, other. Genes in red or green correspond to upregulated or downregulated, respectively. Red lines predict activation; yellow lines indicate inconsistent downstream effect; and gray lines correspond to unpredicted effect. *P < 0.05; **P < 0.01; ns, Not significant.
Mentions: BT-474 cells were exposed to either #hMADS-CM or control medium for the Figure 5D. Proteins were extracted in radioimmunoprecipitation assay buffer and subjected to SDS-PAGE and immunoblot analysis. The antibodies used were anti-phospho-Akt and anti-Akt (Cell Signaling Technology, Beverly, MA, USA) and anti-tubulin (Sigma-Aldrich).

Bottom Line: The results were validated in vivo in a mouse xenograft model.Using a transcriptomic approach, we found that cancer cells undergo major modifications when exposed to adipocyte-conditioned medium.Collectively, our findings underline the importance of adipose tissue in the resistance to trastuzumab and suggest that approaches targeting the adipocyte-cancer cell crosstalk may help sensitize cancer cells to trastuzumab-based therapy.

View Article: PubMed Central - PubMed

Affiliation: Centre de Recherche en Cancérologie de Lyon (CRCL), INSERM UMR 1052, CNRS 5286, 8 Avenue Rockefeller, 69008, Lyon, France. mn.duong@hotmail.com.

ABSTRACT

Introduction: Trastuzumab has been used in the treatment of human epidermal growth factor receptor 2 (HER2)-expressing breast cancer, but its efficacy is limited by de novo or acquired resistance. Although many mechanisms have been proposed to explain resistance to trastuzumab, little is known concerning the role of the tumor microenvironment. Given the importance of antibody-dependent cellular cytotoxicity (ADCC) in the antitumor effect of trastuzumab and the abundance of adipose tissue in the breast, we investigated the impact of adipocytes on ADCC.

Methods: We set up a coculture system to study the effect of adipocytes on ADCC in vitro. The results were validated in vivo in a mouse xenograft model.

Results: We found that adipocytes, as well as preadipocytes, inhibited trastuzumab-mediated ADCC in HER2-expressing breast cancer cells via the secretion of soluble factors. The inhibition of ADCC was not due to titration or degradation of the antibody. We found that adipose cells decreased the secretion of interferon-γ by natural killer cells, but did not alter natural killer cells' cytotoxicity. Preincubation of breast cancer cells with the conditioned medium derived from adipocytes reduced the sensitivity of cancer cells to ADCC. Using a transcriptomic approach, we found that cancer cells undergo major modifications when exposed to adipocyte-conditioned medium. Importantly, breast tumors grafted next to lipomas displayed resistance to trastuzumab in mouse xenograft models.

Conclusions: Collectively, our findings underline the importance of adipose tissue in the resistance to trastuzumab and suggest that approaches targeting the adipocyte-cancer cell crosstalk may help sensitize cancer cells to trastuzumab-based therapy.

No MeSH data available.


Related in: MedlinePlus