Blockade of TGF-β signaling to enhance the antitumor response is accompanied by dysregulation of the functional activity of CD4+CD25+Foxp3+ and CD4+CD25-Foxp3+ T cells

Magdalena J. Polanczyk, Edwin Walker, Daniel Haley, Bella S. Guerrouahen, Emmanuel T. Akporiaye

Research output: Contribution to journalArticle

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Abstract

Background: The pleiotropic cytokine, transforming growth factor (TGF)-β, and CD4+CD25+Foxp3+ regulatory T cells (Tregs) play a critical role in actively suppressing antitumor immune responses. Evidence shows that TGF-β produced by tumor cells promotes tolerance via expansion of Tregs. Our group previously demonstrated that blockade of TGF-β signaling with a small molecule TGF-β receptor I antagonist (SM16) inhibited primary and metastatic tumor growth in a T cell dependent fashion. In the current study, we evaluated the effect of SM16 on Treg generation and function. Methods: Using BALB/c, FoxP3eGFP and Rag-/- mice, we performed FACS analysis to determine if SM16 blocked de novo TGF-β-induced Treg generation in vitro and in vivo. CD4+ T cells from lymph node and spleen were isolated from control mice or mice maintained on SM16 diet, and flow cytometry analysis was used to detect the frequency of CD4+CD25-FoxP3+ and CD4+CD25+FoxP3+ T cells. In vitro suppression assays were used to determine the ability to suppress naive T cell proliferation in vitro of both CD4+CD25+FoxP3+ and CD4+CD25-FoxP3+ T cell sub-populations. We then examined whether SM16 diet exerted an inhibitory effect on primary tumor growth and correlated with changes in FoxP3+expression. ELISA analysis was used to measure IFN-γlevels after 72 h co-culture of CD4+CD25+ T cells from tumor-bearing mice on control or SM16 diet with CD4+CD25- T cells from naive donors. Results: SM16 abrogates TGF-β-induced Treg generation in vitro but does not prevent global homeostatic expansion of CD4+ T cell sub-populations in vivo. Instead, SM16 treatment causes expansion of a population of CD4+CD25-Foxp3+ Treg-like cells without significantly altering the overall frequency of Treg in lymphoreplete naive and tumor-bearing mice. Importantly, both the CD4+CD25-Foxp3+ T cells and the CD4+CD25+Foxp3+ Tregs in mice receiving SM16 diet exhibited diminished ability to suppress naive T cell proliferation in vitro compared to Treg from mice on control diet. Conclusions: These findings suggest that blockade of TGF-β signaling is a potentially useful strategy for blunting Treg function to enhance the anti-tumor response. Our data further suggest that the overall dampening of Treg function may involve the expansion of a quiescent Treg precursor population, which is CD4+CD25-Foxp3+.

Original languageEnglish
Article number219
JournalJournal of translational medicine
Volume17
Issue number1
DOIs
Publication statusPublished - 9 Jul 2019

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T-cells
Transforming Growth Factors
T-Lymphocytes
Nutrition
Tumors
Diet
Bearings (structural)
Neoplasms
Regulatory T-Lymphocytes
Cell proliferation
Population
Cell Proliferation
Growth Factor Receptors
Flow cytometry
Growth
Coculture Techniques
Cell culture
Flow Cytometry
Spleen
Assays

Keywords

  • Anti-tumor response
  • Mice
  • SM16
  • TGF-β
  • Treg subsets

ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology(all)

Cite this

Blockade of TGF-β signaling to enhance the antitumor response is accompanied by dysregulation of the functional activity of CD4+CD25+Foxp3+ and CD4+CD25-Foxp3+ T cells. / Polanczyk, Magdalena J.; Walker, Edwin; Haley, Daniel; Guerrouahen, Bella S.; Akporiaye, Emmanuel T.

In: Journal of translational medicine, Vol. 17, No. 1, 219, 09.07.2019.

Research output: Contribution to journalArticle

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abstract = "Background: The pleiotropic cytokine, transforming growth factor (TGF)-β, and CD4+CD25+Foxp3+ regulatory T cells (Tregs) play a critical role in actively suppressing antitumor immune responses. Evidence shows that TGF-β produced by tumor cells promotes tolerance via expansion of Tregs. Our group previously demonstrated that blockade of TGF-β signaling with a small molecule TGF-β receptor I antagonist (SM16) inhibited primary and metastatic tumor growth in a T cell dependent fashion. In the current study, we evaluated the effect of SM16 on Treg generation and function. Methods: Using BALB/c, FoxP3eGFP and Rag-/- mice, we performed FACS analysis to determine if SM16 blocked de novo TGF-β-induced Treg generation in vitro and in vivo. CD4+ T cells from lymph node and spleen were isolated from control mice or mice maintained on SM16 diet, and flow cytometry analysis was used to detect the frequency of CD4+CD25-FoxP3+ and CD4+CD25+FoxP3+ T cells. In vitro suppression assays were used to determine the ability to suppress naive T cell proliferation in vitro of both CD4+CD25+FoxP3+ and CD4+CD25-FoxP3+ T cell sub-populations. We then examined whether SM16 diet exerted an inhibitory effect on primary tumor growth and correlated with changes in FoxP3+expression. ELISA analysis was used to measure IFN-γlevels after 72 h co-culture of CD4+CD25+ T cells from tumor-bearing mice on control or SM16 diet with CD4+CD25- T cells from naive donors. Results: SM16 abrogates TGF-β-induced Treg generation in vitro but does not prevent global homeostatic expansion of CD4+ T cell sub-populations in vivo. Instead, SM16 treatment causes expansion of a population of CD4+CD25-Foxp3+ Treg-like cells without significantly altering the overall frequency of Treg in lymphoreplete naive and tumor-bearing mice. Importantly, both the CD4+CD25-Foxp3+ T cells and the CD4+CD25+Foxp3+ Tregs in mice receiving SM16 diet exhibited diminished ability to suppress naive T cell proliferation in vitro compared to Treg from mice on control diet. Conclusions: These findings suggest that blockade of TGF-β signaling is a potentially useful strategy for blunting Treg function to enhance the anti-tumor response. Our data further suggest that the overall dampening of Treg function may involve the expansion of a quiescent Treg precursor population, which is CD4+CD25-Foxp3+.",
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AU - Polanczyk, Magdalena J.

AU - Walker, Edwin

AU - Haley, Daniel

AU - Guerrouahen, Bella S.

AU - Akporiaye, Emmanuel T.

PY - 2019/7/9

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N2 - Background: The pleiotropic cytokine, transforming growth factor (TGF)-β, and CD4+CD25+Foxp3+ regulatory T cells (Tregs) play a critical role in actively suppressing antitumor immune responses. Evidence shows that TGF-β produced by tumor cells promotes tolerance via expansion of Tregs. Our group previously demonstrated that blockade of TGF-β signaling with a small molecule TGF-β receptor I antagonist (SM16) inhibited primary and metastatic tumor growth in a T cell dependent fashion. In the current study, we evaluated the effect of SM16 on Treg generation and function. Methods: Using BALB/c, FoxP3eGFP and Rag-/- mice, we performed FACS analysis to determine if SM16 blocked de novo TGF-β-induced Treg generation in vitro and in vivo. CD4+ T cells from lymph node and spleen were isolated from control mice or mice maintained on SM16 diet, and flow cytometry analysis was used to detect the frequency of CD4+CD25-FoxP3+ and CD4+CD25+FoxP3+ T cells. In vitro suppression assays were used to determine the ability to suppress naive T cell proliferation in vitro of both CD4+CD25+FoxP3+ and CD4+CD25-FoxP3+ T cell sub-populations. We then examined whether SM16 diet exerted an inhibitory effect on primary tumor growth and correlated with changes in FoxP3+expression. ELISA analysis was used to measure IFN-γlevels after 72 h co-culture of CD4+CD25+ T cells from tumor-bearing mice on control or SM16 diet with CD4+CD25- T cells from naive donors. Results: SM16 abrogates TGF-β-induced Treg generation in vitro but does not prevent global homeostatic expansion of CD4+ T cell sub-populations in vivo. Instead, SM16 treatment causes expansion of a population of CD4+CD25-Foxp3+ Treg-like cells without significantly altering the overall frequency of Treg in lymphoreplete naive and tumor-bearing mice. Importantly, both the CD4+CD25-Foxp3+ T cells and the CD4+CD25+Foxp3+ Tregs in mice receiving SM16 diet exhibited diminished ability to suppress naive T cell proliferation in vitro compared to Treg from mice on control diet. Conclusions: These findings suggest that blockade of TGF-β signaling is a potentially useful strategy for blunting Treg function to enhance the anti-tumor response. Our data further suggest that the overall dampening of Treg function may involve the expansion of a quiescent Treg precursor population, which is CD4+CD25-Foxp3+.

AB - Background: The pleiotropic cytokine, transforming growth factor (TGF)-β, and CD4+CD25+Foxp3+ regulatory T cells (Tregs) play a critical role in actively suppressing antitumor immune responses. Evidence shows that TGF-β produced by tumor cells promotes tolerance via expansion of Tregs. Our group previously demonstrated that blockade of TGF-β signaling with a small molecule TGF-β receptor I antagonist (SM16) inhibited primary and metastatic tumor growth in a T cell dependent fashion. In the current study, we evaluated the effect of SM16 on Treg generation and function. Methods: Using BALB/c, FoxP3eGFP and Rag-/- mice, we performed FACS analysis to determine if SM16 blocked de novo TGF-β-induced Treg generation in vitro and in vivo. CD4+ T cells from lymph node and spleen were isolated from control mice or mice maintained on SM16 diet, and flow cytometry analysis was used to detect the frequency of CD4+CD25-FoxP3+ and CD4+CD25+FoxP3+ T cells. In vitro suppression assays were used to determine the ability to suppress naive T cell proliferation in vitro of both CD4+CD25+FoxP3+ and CD4+CD25-FoxP3+ T cell sub-populations. We then examined whether SM16 diet exerted an inhibitory effect on primary tumor growth and correlated with changes in FoxP3+expression. ELISA analysis was used to measure IFN-γlevels after 72 h co-culture of CD4+CD25+ T cells from tumor-bearing mice on control or SM16 diet with CD4+CD25- T cells from naive donors. Results: SM16 abrogates TGF-β-induced Treg generation in vitro but does not prevent global homeostatic expansion of CD4+ T cell sub-populations in vivo. Instead, SM16 treatment causes expansion of a population of CD4+CD25-Foxp3+ Treg-like cells without significantly altering the overall frequency of Treg in lymphoreplete naive and tumor-bearing mice. Importantly, both the CD4+CD25-Foxp3+ T cells and the CD4+CD25+Foxp3+ Tregs in mice receiving SM16 diet exhibited diminished ability to suppress naive T cell proliferation in vitro compared to Treg from mice on control diet. Conclusions: These findings suggest that blockade of TGF-β signaling is a potentially useful strategy for blunting Treg function to enhance the anti-tumor response. Our data further suggest that the overall dampening of Treg function may involve the expansion of a quiescent Treg precursor population, which is CD4+CD25-Foxp3+.

KW - Anti-tumor response

KW - Mice

KW - SM16

KW - TGF-β

KW - Treg subsets

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