SUN-300 Neutrophil Extracellular Traps (NETs) Are Present in Tsc2-Deficient Uterine Tumors and May Promote Growth and Metastasis

Abstract Disclosure: M. Wines-Samuelson: None. T. Henson: None. R. Myers: None. S.R. Hammes: None. Background: The relationship between innate immunity and cancer is controversial and currently unclear. Neutrophils, the innate immune cells that rapidly attack infecting pathogens, can directly promote tumor growth by secreting cytokines and other factors that stimulate tumor cell proliferation, and indirectly by secreting factors or expressing cell surface antigens that suppress tumor-targeting T cells. In contrast, neutrophils have been shown to slow tumor growth by presenting neoantigens that stimulate T cell proliferation. To address the role of neutrophils in cancer, our laboratory generated a genetic mouse model (PR-Cre; Tsc2fl/fl, or Tsc2 conditional KO (Tsc2 cKO)) which displays hypertrophic growth of the uterus starting at 18 weeks, with ∼50% of mice developing lung pathology by 30 weeks of age. The tumors in Tsc2 mutant mice resemble the smooth muscle cells tumors present in a rare lung cancer in female patients with lymphangioleiomyelomatosis (LAM), a genetic disease linked to mutations in the human TSC2 gene. Previous work from our lab identified an increase in neutrophils in Tsc2 cKO female mice coinciding with hypertrophic growth of the uterus. Neutrophil depletion, blockade of neutrophil chemotaxis, or inhibition of the enzyme neutrophil elastase, all reduced Tsc2 cKO uterine tumor growth, indicating that neutrophils may be directly promoting tumor progression via neutrophil elastase. We propose that neutrophil extracellular traps (NETs), which contain many factors including neutrophil elastase, are promoting Tsc2 cKO tumor growth. Results: Immunostaining of uterine sections from our Tsc2 cKO mouse revealed significant increases in multiple markers of neutrophil extracellular traps (NETs): myeloperoxidase (MPO), citrullinated histone H3 (H3cit), and extracellular DNA (DAPI). Increased neutrophil NETosis was confirmed by western blotting of uterine lysates from 6 Tsc2 cKO mice aged 18 weeks with anti-MPO and anti-H3cit. Immunofluorescence with in vitro cultures of Tsc2 cKO bone marrow-derived cells independently identified the presence of increased NETs (DAPI+ extracellular DNA + anti-MPO + anti-Ly6G) compared with controls following serum starvation.Conclusions/Future directions: These data support a role for NETosis in uterine tumor progression and/or metastasis to lung. We are currently investigating 1) the contribution of estradiol to NET formation and 2) how tumorigenesis is modulated by NETosis. Presentation: Sunday, July 13, 2025