Abstract PO-017: Targeting murine oral tumors by pharmacological inhibition of b-catenin/CBP epigenetic activity
Authors: Khan Mohammed Muzamil , Reed Eric , Kroehling Lina , Bais Manish , Varelas Xaralabos , Kukuruzinska Maria , Monti Stefano 
Published: September 2023

In previous studies, we have shown that pharmacological blockade of Wnt/β-catenin/CBP activity with small molecule inhibitors is effective in abolishing oncogenic phenotypes in oral squamous cell carcinoma (OSCC). To further characterize changes in Wnt/β-catenin activity during malignant transformation, we used an immunocompetent mouse model of oral cancer induced by a tobacco-derived carcinogen, 4-nitroquinoline-1-oxide (4NQO), which recapitulates the human OSCC mutational landscape and tumor immune environment. We performed single-cell RNA sequencing (scRNAseq) on tongue tissues from healthy mice (n=2), 4NQO-derived mouse OSCC (n=2), and from 4NQO-derived mouse OSCC treated with a pharmacologic grade 𝛽-catenin/CBP inhibitor (n=2). The experiment yielded ~50K cells across all conditions and identified multiple cell types and states, including epithelial and immune cells, as well as endothelia, fibroblasts, and glial cells. We found significant cellular composition changes between 4NQO and 4NQO inhibitor-treated groups, with the proportion of epithelial cells decreasing upon treatment with the 𝛽-catenin/CBP inhibitor, and the endothelial and fibroblast populations increasing in the 𝛽-catenin/CBP inhibitor group. Analysis within the immune compartment showed significant cell type proportion changes between the 4NQO and 4NQO inhibitor-treated groups, including changes in neutrophils population decreasing upon treatment to inhibitor, macrophages, and DCs increasing in the treatment group, along with T- and B-cells. ~50% of the cellular composition in each group comprised of neutrophil population. Further subtyping of the neutrophil population into early and late “neutrotime” subclasses based on published signatures showed enrichment of the early neutrotime class in the inhibitor-treated group and of the late neutrotime class in the 4NQO group. Analysis within the epithelial compartment identified several cell type proportions, including basal (Krt5+, Krt15+), proliferating (Krt5+, Krt14+), and cycling (Top2a, Cdc20) cells increased upon treatment with the inhibitor, and acinar cell-types (Muc5b, Aqp5, Smgc) decreasing upon treatment in comparison with the 4NQO group. The inhibitor-treated group also showed lower proportions of malignant transforming cell states compared to the 4NQO group. A cell-cell communication analysis was performed between epithelial and immune compartments to identify ligand-receptor (LR) interactions indicative of treatment response. Among the significant interactions, the LRs related to collagen organization (Cdh1, Icam1, Lamc1) and major histocompatibility complex genes such as H2-d1, H2-k1, H2-q4, etc., were enriched in the treatment group indicating cellular plasticity and immune regulation processes in comparison with the 4NQO group. Taken together, our results indicate mitigation of cellular heterogeneity and plasticity profiles in 4NQO-induced tumors upon treatment with 𝛽-catenin/CBP inhibitor in OSCC.

Citation Format: Mohammed Muzamil Khan, Eric Reed, Lina Kroehling, Manish Bais, Xaralabos Varelas, Maria Kukuruzinska, Stefano Monti. Targeting murine oral tumors by pharmacological inhibition of b-catenin/CBP epigenetic activity [abstract]. In: Proceedings of the AACR-AHNS Head and Neck Cancer Conference: Innovating through Basic, Clinical, and Translational Research; 2023 Jul 7-8; Montreal, QC, Canada. Philadelphia (PA): AACR; Clin Cancer Res 2023;29(18_Suppl):Abstract nr PO-017.