Abstract A012: Repurposing of the macrolide antibiotic clarithromycin for the prevention of lung cancer

Abstract

Abstract Lung adenocarcinoma (LUAD), particularly K-ras mutant LUAD, is a leading cause of cancer mortality. Therefore, strategies to prevent K-ras-mutant LUAD in its earliest stages in high-risk individuals (e.g., smokers) are urgently needed to reduce the public burden of this fatal disease. We and others have shown that K-ras driven tumorigenesis in lung is intimately linked to chronic inflammation and ultimately, tumor cells immune-escape. The antibiotic clarithromycin (CAM) was identified as one of the most promising candidates for repurposing with demonstrated immunomodulatory and anticancer properties. CAM is widely used and belongs to the macrolide class of antibiotics which are among the safest broad spectrum antimicrobials available. Abundant preclinical and clinical evidence exists demonstrating the in vitro and in vivo anticancer effects of CAM. It has been shown that macrolide antibiotics exert suppression of inflammation without overt immunosuppressive effects mostly through the inhibition of proinflammatory cytokines in vitro and in vivo. In these studies, we tested the lung cancer prevention efficacy of CAM using a Kras mutant lung cancer model. In the CCSPCre; LSL-Kras- G12 D (CC-LR) model, activation of the KrasG12 D mutation takes place in club cells by means of removal of the lox-stop-lox genomic sequence via expression of Cre recombinase under the control of the CCSP promoter. This model is excellent for reproducing the various premalignant to malignant progression steps in the lung. CC-LR mice of both genders were randomly enrolled to four experimental arms comparing three CAM doses: 10mg/kg/day, 50mg/kg/day, and 100mg/kg/day, to vehicle control (H2O). Treatment was administered by oral gavage, 5 times per week for 10 wks., starting at 4 wks. of age. At 14 wks. of age mice were euthanized, lung surface tumors were counted, bronchial lavage fluid, as well as lung samples, were obtained for histological, immunohistochemical (IHC) and qRT-PCR analyses. Clarithromycin treatment led to significant lung cancer prevention efficacy, as determined by lung surface tumor counts. A clear dose response with CAM was observed with a mean lung surface tumor count of 30.2 tumors per mouse for vehicle (n=12 mice), 23.5 for CAM 10mg/kg (n=8), 18.8 for CAM 50mg/kg (n=8) and 13.5 tumors per mouse for the CAM 100mg/kg treatment group (n=12, p=0.0014). A significant decrease in the incidence and multiplicity of premalignant and malignant lesions was also observed in histological analyses. For profiling of the lung immune microenvironment, we analyzed by qRT-PCR the expression of 18 genes identifying various cytokines, cell surface markers and proteins characteristic of specific activation states on the various cell subtypes in the tumor microenvironment. We found significant reduction in the expression of pro-inflammatory cytokines, IL-6, TNF, and IL-1β that are known to have a pro-tumor function in lung tumorigenesis, shown previously by us and other groups. We also found a reduction in the expression of myeloid cell-specific immunosuppressive markers, Arg1 and Fizz 1, that could be due to the reduction in the Gr1+ myeloid cell population, which was detected by means of IHC analyses on the same samples, or due to reprograming of pro-tumor M2 type macrophages toward an anti-tumor M1 phenotype. We also see a trend toward reduction of IL-17 cytokine, which we have previously shown to have an essential role in the promotion of K-ras mutant lung tumors. This was associated with an increase, although not significant, in the expression of anti-tumor Th1-specific transcription factor. However, we see an increase in the expression of PD-1, which could be due to the potential induction of an exhausted T cell phenotype. Taken together, we see a reprograming of the lung microenvironment from a tumor-promoting immunosuppressive phenotype toward an anti-tumor phenotype. This work was supported by NCI PREVENT TORFP75N91019F00131 (TORFP 2019 E-05). Citation Format: Seyed Javad Moghaddam, Tabish Hussain, Melody Zarghooni, Walter V. Velasco, Linda Phan, Michelle I. Savage, Jennifer T. Fox, Shizuko Sei, Powel H. Brown, C. Marcelo Aldaz. Repurposing of the macrolide antibiotic clarithromycin for the prevention of lung cancer [abstract]. In: Proceedings of the Second Biennial NCI Meeting: Translational Advances in Cancer Prevention Agent Development (TACPAD); 2022 Sep 7-9. Philadelphia (PA): AACR; Can Prev Res 2022;15(12 Suppl_2): Abstract nr A012.