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Updated: May 21, 2025
A Novel Feeder-free System for Mass Production of Murine Natural Killer Cells In Vitro
Published on: January 9, 2018
Enhancing human NK cell antitumor function by knocking out SMAD4 to counteract TGFβ and activin A suppression
Anna Rea1, Sara Santana-Hernández2,3, Javier Villanueva1,3
1University Pompeu Fabra (UPF), Barcelona, Spain.
Abstract:
Transforming growth factor beta (TGFβ) and activin A suppress natural killer (NK) cell function and proliferation, limiting the efficacy of adoptive NK cell therapies. Inspired by the partial resistance to TGFβ of NK cells with SMAD4 haploinsufficiency, we used CRISPR-Cas9 for knockout of SMAD4 in human NK cells. Here we show that SMAD4KO NK cells were resistant to TGFβ and activin A inhibition, retaining their cytotoxicity, cytokine secretion and interleukin-2/interleukin-15-driven proliferation. They showed enhanced tumor penetration and tumor growth control, both as monotherapy and in combination with tumor-targeted therapeutic antibodies. Notably, SMAD4KO NK cells outperformed control NK cells treated with a TGFβ inhibitor, underscoring the benefit of maintaining SMAD4-independent TGFβ signaling. SMAD4KO conferred TGFβ resistance across diverse NK cell platforms, including CD19-CAR NK cells, stem cell-derived NK cells and ADAPT-NK cells. These findings position SMAD4 knockout as a versatile and compelling strategy to enhance NK cell antitumor activity, providing a new avenue for improving NK cell-based cancer immunotherapies.
Insights
Genetically modifying natural killer (NK) cells by knocking out SMAD4 enhances their ability to fight cancer. These SMAD4-knockout NK cells resist immune-suppressing factors, improving adoptive cell therapy efficacy.
Area of Science:
- Immunology
- Cell Biology
- Cancer Research
Background:
- Transforming growth factor beta (TGFβ) and activin A inhibit natural killer (NK) cell function and proliferation.
- This suppression limits the effectiveness of adoptive NK cell therapies in cancer treatment.
Purpose of the Study:
- To investigate the potential of SMAD4 knockout to enhance NK cell resistance to TGFβ and activin A.
- To evaluate the therapeutic efficacy of SMAD4-knockout NK cells in preclinical cancer models.
Main Methods:
- CRISPR-Cas9 gene editing was employed to generate SMAD4-knockout (SMAD4KO) human NK cells.
- NK cell function, proliferation, cytotoxicity, and tumor infiltration were assessed in vitro and in vivo.
- The efficacy of SMAD4KO NK cells was compared to control NK cells and NK cells treated with TGFβ inhibitors.
Main Results:
- SMAD4KO NK cells demonstrated resistance to TGFβ and activin A, maintaining cytotoxicity, cytokine secretion, and proliferation.
- These modified NK cells exhibited enhanced tumor penetration and improved tumor growth control as monotherapy and in combination with therapeutic antibodies.
- SMAD4KO NK cells outperformed control NK cells, highlighting the advantage of SMAD4 knockout over transient inhibition.
Conclusions:
- SMAD4 knockout is a robust strategy to confer resistance to TGFβ and activin A in NK cells.
- This approach enhances NK cell antitumor activity across various NK cell platforms, including CAR-NK and stem cell-derived NK cells.
- SMAD4 knockout offers a promising avenue for developing more effective NK cell-based cancer immunotherapies.
