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Background <p>The efficacy of anti-CD19 Chimeric Antigen Receptor (CAR) T-cell therapy has been demonstrated in patients with relapsed or refractory B-cell non-Hodgkin lymphoma (B-NHL). However, therapy-induced senescence (TIS) has been identified as a novel resistance mechanism, potentially exacerbating immunosuppression and impairing CAR-T cell function.</p> Methods <p>Raji and SU-DHL-2 cells were treated with 40 nM doxorubicin (Dox) for 72&#xa0;h to induce TIS, and CCR7 was inhibited with 10&#xa0;µg/mL Cap-100. Cellular senescence was assessed by SA-β-Gal staining and flow cytometry; proliferation and apoptosis by CCK-8 and Annexin V/PI, respectively. CAR-T cytotoxicity was evaluated via flow cytometry. CCR7 expression on T cells from B-NHL patients and healthy controls was measured. Lentiviral transduction regulated CCR7 expression; protein expression and interactions were analyzed by Western blot and Co-IP. NF-κB and ARHGAP/RhoA pathways were inhibited using GS143 and Y27632, respectively.</p> Results <p>CCR7 overexpression enhanced SASP, while knockdown attenuated it. In the Dox-induced TIS model, KMT2D decreased, whereas H3K9me3, CCR7, and LGALS9 increased; CCR7 inhibition reversed these changes. Cap-100 improved CD19 CAR T killing by alleviating exhaustion in co-culture. Mechanistically, Cap-100 stabilized IκBα to inhibit NFκB, and inhibition of NFκB or ROCK reversed pro-senescence. Cap-100 reduced T cell proliferation but did not affect apoptosis or exhaustion. Clinically, CCR7 was lower on T cells from B NHL patients than healthy controls, and higher CCR7 correlated with better T cell quality. Transcriptomics showed Dox activated TIM3/Galectin 9 and PD- 1/PD-L1 pathways, while Dox + Cap-100 suppressed them. Co-IP confirmed interactions among CCR7, KMT2D, and LGALS9. Collectively, Dox induced TIS to upregulate CCR7, increasing SASP and LGALS9, which interacted with TIM-3, causing immune cell exhaustion and reducing killing efficiency; CCR7 blockade reversed this state and enhanced cytotoxicity.</p> CONCLUSIONS <p>Our findings demonstrate that blockade of KMT2D-CCR7-mediated senescence enhances CD19 CAR-T cell anti-tumor activity in B-NHL.</p> Graphical abstract <p></p>

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Targeting CCR7-KMT2D enhances CAR-T cell efficacy by suppressing therapy-induced senescence in B-cell non-Hodgkin lymphoma

  • 期刊论文

  • 出版模式：

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  • 发表日期：

    2026年6月26日

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  • Jinlan Li,
  • Keke Huang,
  • Huiping Wang,
  • Wanjia Chen,
  • Yajie Zhang,
  • Shanyue Jiang,
  • Nengneng Cao,
  • Wanqiu Zhang,
  • Zelin Liu,
  • Jiawen Chen,
  • Dandan Chen,
  • Ziye Yang,
  • Qianshan Tao,
  • Chaohong Liu,
  • Zhimin Zhai,
  • Furun An,
  • Jiyu Wang

摘要

Background

The efficacy of anti-CD19 Chimeric Antigen Receptor (CAR) T-cell therapy has been demonstrated in patients with relapsed or refractory B-cell non-Hodgkin lymphoma (B-NHL). However, therapy-induced senescence (TIS) has been identified as a novel resistance mechanism, potentially exacerbating immunosuppression and impairing CAR-T cell function.

Methods

Raji and SU-DHL-2 cells were treated with 40 nM doxorubicin (Dox) for 72 h to induce TIS, and CCR7 was inhibited with 10 µg/mL Cap-100. Cellular senescence was assessed by SA-β-Gal staining and flow cytometry; proliferation and apoptosis by CCK-8 and Annexin V/PI, respectively. CAR-T cytotoxicity was evaluated via flow cytometry. CCR7 expression on T cells from B-NHL patients and healthy controls was measured. Lentiviral transduction regulated CCR7 expression; protein expression and interactions were analyzed by Western blot and Co-IP. NF-κB and ARHGAP/RhoA pathways were inhibited using GS143 and Y27632, respectively.

Results

CCR7 overexpression enhanced SASP, while knockdown attenuated it. In the Dox-induced TIS model, KMT2D decreased, whereas H3K9me3, CCR7, and LGALS9 increased; CCR7 inhibition reversed these changes. Cap-100 improved CD19 CAR T killing by alleviating exhaustion in co-culture. Mechanistically, Cap-100 stabilized IκBα to inhibit NFκB, and inhibition of NFκB or ROCK reversed pro-senescence. Cap-100 reduced T cell proliferation but did not affect apoptosis or exhaustion. Clinically, CCR7 was lower on T cells from B NHL patients than healthy controls, and higher CCR7 correlated with better T cell quality. Transcriptomics showed Dox activated TIM3/Galectin 9 and PD- 1/PD-L1 pathways, while Dox + Cap-100 suppressed them. Co-IP confirmed interactions among CCR7, KMT2D, and LGALS9. Collectively, Dox induced TIS to upregulate CCR7, increasing SASP and LGALS9, which interacted with TIM-3, causing immune cell exhaustion and reducing killing efficiency; CCR7 blockade reversed this state and enhanced cytotoxicity.

CONCLUSIONS

Our findings demonstrate that blockade of KMT2D-CCR7-mediated senescence enhances CD19 CAR-T cell anti-tumor activity in B-NHL.

Graphical abstract