Neutrophil extracellular traps induced by chemotherapy inhibit tumor growth in murine models of colorectal cancer
Yamu Li, Sulin Wu, Yiqing Zhao, Trang Dinh, Dongxu Jiang, J. Eva Selfridge, George Myers, Yuxiang Wang, Xuan Zhao, Suzanne Tomchuck, George Dubyak, Richard T. Lee, Bassam Estfan, Marc Shapiro, Suneel Kamath, Amr Mohamed, Stanley Ching-Cheng Huang, Alex Y. Huang, Ronald Conlon, Smitha Krishnamurthi, Jennifer Eads, Joseph E. Willis, Alok A. Khorana, David Bajor, Zhenghe Wang
Yamu Li, Sulin Wu, Yiqing Zhao, Trang Dinh, Dongxu Jiang, J. Eva Selfridge, George Myers, Yuxiang Wang, Xuan Zhao, Suzanne Tomchuck, George Dubyak, Richard T. Lee, Bassam Estfan, Marc Shapiro, Suneel Kamath, Amr Mohamed, Stanley Ching-Cheng Huang, Alex Y. Huang, Ronald Conlon, Smitha Krishnamurthi, Jennifer Eads, Joseph E. Willis, Alok A. Khorana, David Bajor, Zhenghe Wang
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Research Article Oncology

Neutrophil extracellular traps induced by chemotherapy inhibit tumor growth in murine models of colorectal cancer

Abstract

Neutrophil extracellular traps (NETs), a web-like structure of cytosolic and granule proteins assembled on decondensed chromatin, kill pathogens and cause tissue damage in diseases. Whether NETs can kill cancer cells is unexplored. Here, we report that a combination of glutaminase inhibitor CB-839 and 5-FU inhibited the growth of PIK3CA-mutant colorectal cancers (CRCs) in xenograft, syngeneic, and genetically engineered mouse models in part through NETs. Disruption of NETs by either DNase I treatment or depletion of neutrophils in CRCs attenuated the efficacy of the drug combination. Moreover, NETs were present in tumor biopsies from patients treated with the drug combination in a phase II clinical trial. Increased NET levels in tumors were associated with longer progression-free survival. Mechanistically, the drug combination induced the expression of IL-8 preferentially in PIK3CA-mutant CRCs to attract neutrophils into the tumors. Further, the drug combination increased the levels of ROS in neutrophils, thereby inducing NETs. Cathepsin G (CTSG), a serine protease localized in NETs, entered CRC cells through the RAGE cell surface protein. The internalized CTSG cleaved 14-3-3 proteins, released BAX, and triggered apoptosis in CRC cells. Thus, our studies illuminate a previously unrecognized mechanism by which chemotherapy-induced NETs kill cancer cells.

Authors

Yamu Li, Sulin Wu, Yiqing Zhao, Trang Dinh, Dongxu Jiang, J. Eva Selfridge, George Myers, Yuxiang Wang, Xuan Zhao, Suzanne Tomchuck, George Dubyak, Richard T. Lee, Bassam Estfan, Marc Shapiro, Suneel Kamath, Amr Mohamed, Stanley Ching-Cheng Huang, Alex Y. Huang, Ronald Conlon, Smitha Krishnamurthi, Jennifer Eads, Joseph E. Willis, Alok A. Khorana, David Bajor, Zhenghe Wang

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Figure 5

CTSG in NETs inhibits tumor growth.

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CTSG in NETs inhibits tumor growth. (A–C) The HCT116 CRC cells were trea...
(A–C) The HCT116 CRC cells were treated with the indicated recombinant proteins (10 μg/mL) overnight. Cell lysates were blotted with the indicated antibodies (A). The HCT116 cells were treated with the indicated recombinant proteins. Cells were treated with camptothecin (3 μM) in parallel as a positive control. Caspase 3 activities were measured (B) (n = 3/group). Cell numbers are shown in C (n = 3/group). (D–G) HCT116 cells were treated with indicated concentrations of recombinant CTSG. Caspase 3 activities are shown in D (n = 3/group), cell numbers were counted in E (n = 3/group). Annexin V staining and quantifications are shown in F and G (n = 3/group). (H–J) The indicated xenograft tumors were established in nude mice and treated with vehicle or the drug combination with or without CTSGi (intratumor injection, 5 mice/group). (K–N) Tunel staining of the tumors is shown in H–J. Representative images of HCT116 tumors are shown in K. Quantifications are shown in L–N (n = 15/group). (O–Q) Tumors from H stained with anti-MPO and anti-H3cit antibodies. Representative images of HCT116 tumors are shown in O, and quantifications are shown in P and Q (n = 15/group). Scale bar: 50 μm. 1-way ANOVA (B–E, G, L–N, P, and Q) or 2-way ANOVA (H–J) was used for statistical analysis. *P < 0.05; **P < 0.01; ***P < 0.001; ****P < 0.0001.