American Association for Cancer Research
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FIGURE 4 from Epithelial Expressed B7-H4 Drives Differential Immunotherapy Response in Murine and Human Breast Cancer

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posted on 2024-04-24, 14:20 authored by Elizabeth C. Wescott, Xiaopeng Sun, Paula Gonzalez-Ericsson, Ann Hanna, Brandie C. Taylor, Violeta Sanchez, Juliana Bronzini, Susan R. Opalenik, Melinda E. Sanders, Julia Wulfkuhle, Rosa I. Gallagher, Henry Gomez, Claudine Isaacs, Vijaya Bharti, John T. Wilson, Tarah J. Ballinger, Cesar A. Santa-Maria, Payal D. Shah, Elizabeth C. Dees, Brian D. Lehmann, Vandana G. Abramson, Gillian L. Hirst, Lamorna Brown Swigart, Laura J. van ˈt Veer, Laura J. Esserman, Emanuel F. Petricoin, Jennifer A. Pietenpol, Justin M. Balko

Overexpression of B7-H4 in the EMT6 murine model induced resistance to anti-PD-L1 ICI. A, EMT6 cells were virally transduced with the pBabe-B7-H4 retroviral vector and overexpress exogenous murine B7-H4. B, EMT6 B7-H4+ tumors maintain high B7-H4 expression in vivo assessed by IHC. C, Animals were orthotopically injected with EMT6 cells ± B7-H4 and treated 1x per week with anti-PD-L1 at 200 µg (first dose) or 100 µg (subsequent doses) for 4 weeks, after tumors reached 100 mm3. D, EMT6-B7-H4+ tumors are significantly resistant to anti-PD-L1 immunotherapy compared with control tumors. Data were analyzed by one-way ANOVA of individual AUC values with Tukey post hoc test for multiple comparisons between EMT6 anti-PD-L1 and EMT6-B7-H4+ anti-PD-L1–treated groups (P = 0.0174, EMT6 Isotype n = 21, EMT6 anti-PD-L1 n = 23, EMT6-B7-H4+ Isotype n = 21, EMT6-B7-H4+ anti-PD-L1 n = 23. Data were collected from a total of three independent experiments). E and F, When tumor response is categorized into three groups, EMT6-B7-H4+ tumors have overall greater intrinsic resistance to treatment and reduced complete response compared with EMT6 control tumors (P = 0.035, χ2 = 6.683, df = 2, n = 23 mice for EMT6 parental tumors and n = 31 mice for B7-H4 tumors). G, EMT6-B7-H4+ tumors were stained by mIF. B7-H4 is expressed on CD45 tumor cells in vivo. Representative image shown. Scale bar 10 µm. H, Quantification of G, n = 3 mice. Data analyzed by unpaired t test. Data were analyzed in GraphPad Prism v10.


HHS | NIH | National Cancer Institute (NCI)

U.S. Department of Defense (DOD)



Combinations of immune checkpoint inhibitors (ICI, including anti-PD-1/PD-L1) and chemotherapy have been FDA approved for metastatic and early-stage triple-negative breast cancer (TNBC), but most patients do not benefit. B7-H4 is a B7 family ligand with proposed immunosuppressive functions being explored as a cancer immunotherapy target and may be associated with anti-PD-L1 resistance. However, little is known about its regulation and effect on immune cell function in breast cancers. We assessed murine and human breast cancer cells to identify regulation mechanisms of B7-H4 in vitro. We used an immunocompetent anti-PD-L1–sensitive orthotopic mammary cancer model and induced ectopic expression of B7-H4. We assessed therapy response and transcriptional changes at baseline and under treatment with anti-PD-L1. We observed B7-H4 was highly associated with epithelial cell status and transcription factors and found to be regulated by PI3K activity. EMT6 tumors with cell-surface B7-H4 expression were more resistant to immunotherapy. In addition, tumor-infiltrating immune cells had reduced immune activation signaling based on transcriptomic analysis. Paradoxically, in human breast cancer, B7-H4 expression was associated with survival benefit for patients with metastatic TNBC treated with carboplatin plus anti-PD-L1 and was associated with no change in response or survival for patients with early breast cancer receiving chemotherapy plus anti-PD-1. While B7-H4 induces tumor resistance to anti-PD-L1 in murine models, there are alternative mechanisms of signaling and function in human cancers. In addition, the strong correlation of B7-H4 to epithelial cell markers suggests a potential regulatory mechanism of B7-H4 independent of PD-L1. This translational study confirms the association of B7-H4 expression with a cold immune microenvironment in breast cancer and offers preclinical studies demonstrating a potential role for B7-H4 in suppressing response to checkpoint therapy. However, analysis of two clinical trials with checkpoint inhibitors in the early and metastatic settings argue against B7-H4 as being a mechanism of clinical resistance to checkpoints, with clear implications for its candidacy as a therapeutic target.