Lung cancer is one of the deadliest and most common forms of cancer globally, with 80-85% of cases being non-small cell lung cancer (NSCLC). Treatment for NSCLC includes immune checkpoint inhibitors, such as anti-PD-1 therapy. Anti-PD-1 blocks the interaction between PD-1 and PD-L1, leading to enhanced anti-tumor responses. Although anti-PD-1 therapy leads to clinical benefit in a subset of NSCLC patients, many patients harboring mutations in Liver Kinase 1 (LKB1/STK11) show resistance to anti-PD-1, which is associated with dominance of myeloid derived suppressor cells (MDSCs) in the tumor microenvironment (TME). MDSCs are a heterogeneous population of immature myeloid cells that suppress T cell function and promote tumor growth. Our previous studies demonstrate that all-trans retinoic acid (ATRA), an FDA-approved drug, exerts anti-tumor efficacy against LKB1-deficient murine NSCLC tumors by inhibiting the suppressive function of MDSC. We further demonstrate that combination therapy of anti-PD-1 and ATRA promotes T cell proliferation and improves anti-PD-1 efficacy. Our preliminary data reveal elevated levels of reactive oxygen species (ROS) in MDSCs from tumor bearing mice treated with ATRA, which may cause mitochondrial instability and activation of a type 1 interferon (IFN) response. We hypothesize that ATRA inhibits the suppressive function of MDSC by stimulating production of type 1 IFN from MDSCs in the TME. To test this hypothesis, we propose to perform ATRA treatment in murine NSCLC models, followed by tumor processing and MDSC isolation to analyze for levels of Interferon-beta 1 (IFN-b1) and downstream targets of type 1 IFN pathway via qPCR and ELISA.

Manipulating the activation of immune cells to promote antitumor immunity has shown great promise and has resulted in newly developed immunotherapies. One molecule of interest for use as an immunotherapy agent is creatine. Previous studies demonstrated that knockout of the CrT gene (Slc6a8), which encodes a cell surface creatine transporter, decreased creatine uptake into the cell, which resulted in decreased antitumor immunity. They also show that creatine supplementation enhanced antitumor immunity by providing energy for T cell functions. This function of creatine as a metabolic regulator suggests that creatine may also increase antitumor immunity through the metabolic activation of other immune cells, such as dendritic cells. We show that creatine supplementation of Slc6a8 WT mouse bone marrow-derived dendritic cells (BMDCs) increases the amount of ATP available for use. We also show that this increased source of ATP activates the c-Jun/Ap-1 pathway in BMDCs in vitro. This can result in increased expression of chemokines and CD40 costimulatory proteins, enhanced BMDC viability, and CD8+ T cell enhancement. Our results suggest the potential use of creatine supplementation as a way to enhance dendritic cell antitumor immunotherapies.

Aging is the biggest risk factor for multiple diseases including cancer, neurodegeneration, and cardiovascular disease, amongst others, which calls for an increase in care and treatments for the growing aging population. Cellular senescence, caused by cellular damage that leads to cell cycle arrest, is thought to contribute to the aging process and is characterized by the secretion of pro-inflammatory molecules known as the Senescent Associated Secretory Phenotype (SASP). Macrophages are ubiquitous innate-immune cells that serve multiple functions including protection from pathogens, maintaining tissue homeostasis, and function to control tissue metabolism. Our lab has shown that macrophages can be induced to become senescent using DNA damaging stimuli such as irradiation and chemotherapeutic drugs. Our data suggest macrophages may represent a key source of senescent cells in aging tissues, however, the molecular mechanisms driving senescence in macrophages and their link to age-related diseases remain unclear. In our study, we aim to determine which genes contribute to cellular senescence, with a focus on the cell cycle regulator, Cyclin D2 (CcnD2), as our lab has shown its expression is significantly increased in senescent cells. Using macrophages derived from mouse bone marrow, we use Ccnd2 knockout mice and genetic editing using CRISPR-Cas9 to investigate how Cyclin D2 regulates SASP-related genes such as NF-kB, IL-6, and TNF-a. Understanding the molecular mechanisms regulating macrophage senescence may provide insight into treating age-related diseases.

Fusobacterium nucleatum is a Gram-negative anaerobic oral commensal that can be pathogenic when it spreads to extraoral sites. Motility, a common virulence factor, has been previously thought to be absent in F. nucleatum, but conflicting data has created a gap in knowledge. Using biological assays and live cell microscopy, we have observed motility in F. nucleatum at both the colony (macro) and single bacterium (micro) scales. In the macroscale assay, fusobacterial motility was observed via colony expansion on semisolid surfaces, whereas in the microscale assay, the movement of single bacteria in an anaerobic chamber was monitored under a microscope. Bacterial movement was quantified using R and ImageJ. Since F. nucleatum does not have flagella, we decided to investigate alternate mechanisms of motility, such as lipopolysaccharide (LPS)-mediated motility as observed in other Gram-negative bacteria. Using a sequence-defined mutant library developed in the lab, we identified many LPS mutants that were severely defective in motility in both the macro and microscale assays. Since motility is critical for bacterial survival and virulence, evidence for motility in F. nucleatum may be a possible mechanism for pathogenesis in extraoral sites, which will be further investigated.