Entinostat induces antitumor immune responses through immune editing of tumor neoantigens
Andrew S. Truong, Mi Zhou, Bhavani Krishnan, Takanobu Utsumi, Ujjawal Manocha, Kyle G. Stewart, Wolfgang Beck, Tracy L. Rose, Matthew I. Milowsky, Xiaping He, Christof C. Smith, Lisa M. Bixby, Charles M. Perou, Sara E. Wobker, Sean T. Bailey, Benjamin G. Vincent, William Y. Kim
Andrew S. Truong, Mi Zhou, Bhavani Krishnan, Takanobu Utsumi, Ujjawal Manocha, Kyle G. Stewart, Wolfgang Beck, Tracy L. Rose, Matthew I. Milowsky, Xiaping He, Christof C. Smith, Lisa M. Bixby, Charles M. Perou, Sara E. Wobker, Sean T. Bailey, Benjamin G. Vincent, William Y. Kim
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Research Article Oncology

Entinostat induces antitumor immune responses through immune editing of tumor neoantigens

Abstract

Although immune-checkpoint inhibitors (ICIs) have been a remarkable advancement in bladder cancer treatment, the response rate to single-agent ICIs remains suboptimal. There has been substantial interest in the use of epigenetic agents to enhance ICI efficacy, although precisely how these agents potentiate ICI response has not been fully elucidated. We identified entinostat, a selective HDAC1/3 inhibitor, as a potent antitumor agent in our immune-competent bladder cancer mouse models (BBN963 and BBN966). We demonstrate that entinostat selectively promoted immune editing of tumor neoantigens, effectively remodeling the tumor immune microenvironment, resulting in a robust antitumor response that was cell autonomous, dependent upon antigen presentation, and associated with increased numbers of neoantigen-specific T cells. Finally, combination treatment with anti–PD-1 and entinostat led to complete responses and conferred long-term immunologic memory. Our work defines a tumor cell–autonomous mechanism of action for entinostat and a strong preclinical rationale for the combined use of entinostat and PD-1 blockade in bladder cancer.

Authors

Andrew S. Truong, Mi Zhou, Bhavani Krishnan, Takanobu Utsumi, Ujjawal Manocha, Kyle G. Stewart, Wolfgang Beck, Tracy L. Rose, Matthew I. Milowsky, Xiaping He, Christof C. Smith, Lisa M. Bixby, Charles M. Perou, Sara E. Wobker, Sean T. Bailey, Benjamin G. Vincent, William Y. Kim

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Figure 5

B2m CRISPR partially rescues the antitumor effect of entinostat.

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B2m CRISPR partially rescues the antitumor effect of entinostat. (A) Im...
(A) Immunoblot of BBN963 cells with B2m knockout using 9 different individual sgRNA constructs. (B) Flow cytometry graph of H-2Kb (class 1 MHC) cell-surface expression in B2m-knockout BBN963 cells in A with sgRNA no. 4 and no. 5. (C) Average volume of BBN963_EV (empty vector) and BBN963_sgB2m (B2m knockout) tumors in response to entinostat (12 mpk) in C57BL/6 mice. n = 7–13 mice per group. Significance was calculated using 2-way ANOVA followed by Tukey’s multiple-comparison test. (D) Waterfall plot of individual tumor volumes in C at end points relative to pretreatment baseline. End points were tumor burden and ulceration. (E) Average volume of BBN966_EV (empty vector) and BBN966_sgB2m (B2m knockout) tumors in response to entinostat (12 mpk) in C57BL/6 mice. n = 5–6 mice per group. Significance was calculated using 2-way ANOVA followed by Tukey’s multiple-comparison test. (F) Waterfall plot of individual tumor volumes in E at end points relative to pretreatment baseline. End points were tumor burden and ulceration (G) Schematic of how the in vitro T cell killing assays were set up. (H) Immunoblots of BBN963 cells treated with various concentrations of entinostat for 24 hours and blotted for the indicated antibodies. (I) Bar graph showing the result of T cell killing assays. CellTiter-Glo was used to quantify viable BBN963 cells at the end of the 72-hour coculture. Significance was calculated using t test. Data are represented as mean ± SD. ***P < 0.001; ****P < 0.0001.