Abstract 6344: Effects of psychological stress early in NNK-induced lung carcinogenesis in young female mice

Abstract Tobacco smoke contains many known carcinogens, chief among which is nicotine-derived nitrosamine ketone (NNK). However, even among individuals with high exposure levels, only a minority develop lung cancer. Prior research highlights additional exogenous (e.g., air pollution) and endogenous (e.g., estrogen exposure) factors that may increase risk. Currently lacking in the literature is consideration of possible effects of psychological stress. To experimentally test stress effects, we utilized a well-established mouse lung adenocarcinoma model with NNK exposure of young female mice, along with a mouse stress protocol. We previously reported that repeated exposure to stress before, during and after 4 weeks of exposure to NNK (IP; 2x/week; 3 mg/injection) increased both overall lesion size and the total number of lung lesions compared to controls when assessed 19 weeks post completion of NNK exposure. The present study used the same protocol to test stress effects earlier in the process of NNK carcinogenesis. FVB/N female mice (4-6 wk) bought from a commercial supplier were acclimated for two weeks before study initiation. The repeated stress protocol was then begun (Week-0) for the Stress group (n=40) and continued throughout the study, while the Control group (n=40) remained in their home cages. The stress protocol included social disruption via changes in cage mates at the time of routine cage cleaning (2x/week) and 90-minute restraint stress prior to a cage mate change (1x/week). NNK treatment began on week-1 (IP; 2x/week; 3 mg/injection) and continued for 4 weeks. At 8 and 15 weeks after the end of NNK exposure, 20 mice in each study group were euthanized. Lungs were formalin inflated. Surface lung lesions were counted under a dissecting microscope and lesion sizes were objectively measured (blind) using Motic Images software. Lungs from mice euthanized at 15 weeks were then imbedded in paraffin and serially sectioned (4 µm) to allow H&E confirmation of lesion numbers and sizes by a veterinary pathologist and to provide slides for ongoing immunohistochemical characterization. Statistical analysis (ANOVA) revealed significant main effects of study group, with lungs from stressed mice having larger overall lesion sizes (p=.007), and total number of lesions (p=.012), compared to controls. As expected, those two outcomes were also significantly increased in the lungs from mice euthanized at 15- vs. 8-weeks (no interaction with group). Providing independent corroboration of the Motic imaging results, data from the histological evaluation of lesion sizes and numbers were significantly correlated (p's <.001). Conclusion: We replicated and extended our initial study demonstrating effects of repeated stress in an NNK model of lung cancer in young female mice to earlier phases of NNK-induced lung carcinogenesis. Citation Format: Dana H. Bovbjerg, Laura P. Stabile, Beatriz Kanterewicz, Jessica Manculich, Jessie R. Nedrow, Patrick Tarwater, Lora H. Rigatti, Frank J. Jenkins. Effects of psychological stress early in NNK-induced lung carcinogenesis in young female mice [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 6344.