In situ expansion and reprogramming of Kupffer cells elicit potent tumoricidal immunity against liver metastasis
Wei Liu, … , Lu Li, Zhutian Zeng
Wei Liu, … , Lu Li, Zhutian Zeng
Published February 23, 2023
Citation Information: J Clin Invest. 2023;133(8):e157937. https://doi.org/10.1172/JCI157937.
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Research Article Hepatology Immunology

In situ expansion and reprogramming of Kupffer cells elicit potent tumoricidal immunity against liver metastasis

Abstract

Liver metastasis represents one of the most frequent malignant diseases with no effective treatment. Functional reprogramming of Kupffer cells (KCs), the largest population of hepatic macrophages, holds promise for treating liver cancer, but remains seldom exploited. Taking advantage of the superior capacity of KCs to capture circulating bacteria, we report that a single administration of attenuated Escherichia coli producing clustered regularly interspersed short palindromic repeats CasΦ (CRISPR/CasΦ) machinery enables efficient editing of genes of interest in KCs. Using intravital microscopy, we observed a failure of tumor control by KCs at the late stage of liver metastasis due to KC loss preferentially in the tumor core and periphery, resulting in inaccessibility of these highly phagocytic macrophages to cancer cells. Simultaneous disruption of MafB and c-Maf expression using the aforementioned engineered bacteria could overcome KC dysfunction and elicit remarkable curative effects against several types of metastatic liver cancer in mice. Mechanistically, bacterial treatment induced massive proliferation and functional reprogramming of KCs. These cells infiltrated into the tumor, dismantled macrometastases by nibbling cancer cells, and skewed toward proinflammatory macrophages to unleash antitumor T cell responses. These findings provide an immunotherapy strategy that could be applicable for treating liver metastasis and highlight the therapeutic potential of targeting tissue-resident macrophages in cancer.

Authors

Wei Liu, Xia Zhou, Qi Yao, Chen Chen, Qing Zhang, Keshuo Ding, Lu Li, Zhutian Zeng

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Figure 2

Genetic modification of KCs in situ via bacterial delivery of CRISPR machinery.

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Genetic modification of KCs in situ via bacterial delivery of CRISPR mac...
(A) Intravital liver images at day 7 after injection with 1 × 109 CFU E. coli TOP10 carrying either the pX459-2U6 backbone vector or pX459-2U6-2sgCrig plasmids. Scale bars: 200 μm. Original magnification, zoomed images: ×7.5. (B) Normalized fluorescence intensity of CRIg per FOV. (C) Survival of mice infected with 1 × 109 CFU E. coli TOP10 (n = 108) or ClearColi (n = 92); pooled data are shown. (D) Normalized hepatic mRNA levels of Ccl2, Tnfa, and Il6 and (E) serum levels of ALT and AST at 24 hours after injection with 1 × 109 CFU of TOP10 or ClearColi. (F) Intravital liver images and (G) normalized fluorescence intensity of CRIg per FOV at day 7 after injection with ClearColi harboring either the Crig-targeting CRISPR/Cas9 or CRISPR/CasΦ plasmids. Scale bars: 50 μm. Original magnification, zoomed images: ×4.0. (H) Representative flow cytometric plot of KCs at days 0 and 7 after ClearColi-sgRosa26 (1 × 109 CFU) injection. (I) Quantification of the percentages of F4/80+TIM4+ resident KCs. Fate-mapping experiments showing the percentages of tdTomato-labeled KCs in (J) Clec4f-CreER: RCL-tdT or (K) Ccr2-CreER: RCL-tdT mice at days 0 and 7 after ClearColi-sgCrig injection. For B and G, each circle represents 1 FOV, and a total of 30 randomly selected FOVs from 4 mice per group were analyzed. For D, E, and I–K, each circle represents 1 mouse. Representative or pooled data from at least 2 independent experiments are shown. Data are represented as mean ± SEM. **P < 0.01; ***P < 0.001, unpaired Student’s t test (B, G, I–K); 1-way ANOVA with Tukey’s test (D and E); 2-sided log-rank test (C).