Low-dose statins restore innate immune response in breast cancer cells via suppression of mutant p53

Abstract

Background: Breast cancer's high recurrence and treatment side effects demand safer, more effective therapies. Mutations in the critical TP53 gene, which normally prevents cancer, can instead promote it. This study explores if low-dose statins can curb mutant p53 activation in breast cancer's immune signaling, hindering tumor immune evasion.

Methods: The study used diverse breast cancer cell lines with varying p53 statuses. Techniques included Western blot, transfection, qRT-PCR, co-immunoprecipitation, nuclear fractionation, and immunohistochemistry. In vivo experiments used BALB/c mice, with bioinformatics analysis via cBioPortal.

Results: The study found that suppressing mutant p53 restores innate immunity and enhances cancer treatment. Low-dose statins promoted IRF3 nuclear translocation by inhibiting mutant p53. Lovastatin treatment in vivo increased phosphorylated TBK1 and IRF3 levels and induced CD8⁺ T lymphocyte infiltration in tumors.

Conclusion: The findings suggest low-dose statins can enhance innate immunity in breast cancer by degrading mutant p53, offering new treatment possibilities. Caution is advised, and further research is needed to address limitations and provide solid evidence for clinical use.

Keywords: CGAS; STING; breast cancer; p53; statin.

FIGURE 1

Mutant p53 inhibits innate immune signaling in breast cancer. Survival analysis of TP53-Mutant and TP53-NonMutant groups in breast cancer patients (A). Expression of STING in breast cancer with TP53-Mutant group versus TP53-NonMutant group, *p < 0.05 (B). Immunoprecipitation of mutant P53 in whole-cell lysates from MDA-MB-231 (p53^R280K) and Sk-Br-3 (p53^R175H) cells. Immunoblot analysis of lysates and immunoprecipitates (C). Western blot analysis of cGAS-STING pathway proteins in the human breast cancer cell lines MDA-MB-231 (P53^R280K) and Sk-Br-3 (P53^R175H) following shp53 (D). Western blot analysis of cGAS-STING pathway proteins in 4T1 cells transfected with p53R280K, using 4T1 (p53^null) cells transfected with shCOO2 as the control (E).

FIGURE 2

Prolonged low-dose statins reduce mutant p53 and activate innate immunity in breast cancer cells. Western blot analysis of MDA-MB-231 (p53^R280K) and 4T1 (p53^R280K) cells treated with low-dose statins collected at 0, 24, 48, 72, 96, and 120 h (A). MDA-MB-231 (p53^R280K) cells were treated with indicated concentrations of fluvastatin (0, 0.12, 0.25, 0.5 μM) and lovastatin (0, 4 μM) for 120 h, respectively, and cell samples were collected for western blot experiments (B). Empty vector plasmid shC002 transfected 4T1 (p53^null) cells and mutant p53-transfected 4T1 (p53^R280K) cells were treated with indicated concentrations of fluvastatin (0, 0.5 μM) and lovastatin (0, 0.5 μM) for 120 h, respectively, and cell samples were collected for western blot experiments (C, D).

FIGURE 3

Low-dose statins boost IRF3 nuclear translocation and ISGs in mutant p53 breast cancer. MDA-MB-231 (p53R280K) cells were treated with indicated concentrations of fluvastatin (0, 0.12, 0.25, 0.5 μM) for 120 h, followed by nuclear-cytoplasmic fractionation experiments and western blot analysis to assess IRF3 protein expression (A). 4T1 (p53^null) cells transfected with empty vector plasmid shC002 and mutant p53-transfected 4T1 (p53^R280K) cells were treated with indicated concentrations of fluvastatin (0, 0.5 μM) for 120 h, then subjected to nuclear-cytoplasmic fractionation experiments and western blot analysis to evaluate IRF3 protein expression (B). MDA-MB-231 (p53^R280K) cells were treated with 0.5 μM fluvastatin and 0.5 μM lovastatin for 120 h, respectively, followed by cell sample collection for RT-PCR detection of CXCL10 and ISG15 mRNA. N = 3, *p < 0.05, **p < 0.01, ***p < 0.001 (C). Mutant p53-transfected 4T1 (p53^R280K) cells were treated with 0.5 μM fluvastatin and 0.5 μM lovastatin for 120 h, respectively, and cell samples were collected for RT-PCR analysis of IFNB1 mRNA. N = 3, **p < 0.01 (D).

FIGURE 4

STING gene knockdown reduces low-dose statin-induced innate immunity activation. MDA-MB-231 cells transfected with shC002, shSTING-1, and shSTING-2 were treated with 0.5 μM lovastatin for 120 h followed by western blot experiments (A). MDA-MB-231 cells transfected with shC002, shSTING-1, and shSTING-2 were treated with 0.5 μM lovastatin for 120 h followed by RT-PCR experiments. N = 3, *p < 0.05, **p < 0.01 (B).

FIGURE 5

Statin inhibition of proliferation in mutant p53 breast tumors in an intact host immune system. Monitoring of body weight during a 3-week treatment period of 4T1 (p53^R280K) lovastatin and control groups. 4T1 (p53^R280K) treated group n = 9, 4T1 (p53^R280K) control group n = 10. *p < 0.05 (A). Monitoring of mammary tumor growth in the two groups mentioned in (A) over the 3-week treatment period. *p < 0.05 (B). Photographic documentation of tumors from dissected BALB/c mice in the two groups mentioned in (A) following treatment termination (C). Statistical analysis of tumor volumes from (C), *p < 0.05 (D). Statistical analysis of tumor weights from (C), *p < 0.05 (E).

FIGURE 6

Statin activation of innate immune suppression in mutant p53 breast cancer tumor growth. 4T1 (p53^R280K) BALB/c mice were euthanized on day 23, tumors were excised, sectioned, and subjected to immunoblot analysis of tumor tissue (A). Representative immunohistochemistry images of p53, CD3⁺CD8⁺ T lymphocyte infiltration. Magnification is 200* (B).