A BSTRACT Introduction: How fluoride (F – ) protects dental enamel from caries is here conveyed to dental health-care providers by making simplifying approximations that accurately convey the essential principles, without obscuring them in a myriad of qualifications. Materials and Methods: We approximate that dental enamel is composed of calcium hydroxyapatite (HAP), a sparingly soluble ionic solid with the chemical formula Ca 10 (PO 4 ) 6 (OH) 2 . Results: The electrostatic forces binding ionic solids together are described by Coulomb’s law, which shows that attractions between opposite charges increase greatly as their separation decreases. Relatively large phosphate ions (PO 4 3– ) dominate the structure of HAP, which approximates a hexagonal close-packed structure. The smaller Ca 2+ and OH – ions fit into the small spaces (interstices) between phosphates, slightly expanding the close-packed structure. F – ions are smaller than OH – ions, so substituting F – for OH – allows packing the same number of ions into a smaller volume, increasing their forces of attraction. Dental decay results from tipping the solubility equilibrium Ca 10 (PO 4 ) 6 (OH) 2 (s) ⇔ 10Ca 2+ (aq) + 6PO 4 2– (aq) + 2OH – (aq) toward dissolution. HAP dissolves when the product of its ion concentrations, [Ca 2+ ] 10 ×[PO 4 3– ] 6 ×[OH – ] 2 , falls below the solubility product constant (Ksp) for HAP. Conclusion: Because of its more compact crystal structure, the Ksp for fluorapatite (FAP) is lower than the Ksp for HAP, so its ion product, [Ca 2+ ] 10 ×[PO 4 3– ] 6 ×[F – ] 2 , must fall further before demineralization can occur. Lowering the pH of the fluid surrounding enamel greatly reduces [PO 4 3– ] (lowering the ion products of HAP and FAP equally), but [OH – ] falls much more rapidly than [F – ], so FAP better resists acid attack.

Cancer development and progression is associated with metabolic reprogramming of tumor cells required to meet their proliferative, bioenergetic and survival challenges. Some metabolites, including a non-essential amino acid, serine, have been shown to play a role in tumorigenesis by promoting aggressive cell states, such as cancer stem cells (CSCs). Previous studies from our laboratory showed that the nuclear branch of the Wnt/β-catenin signaling pathway, the β-catenin/CBP/MLL1 axis, enhanced CSC states during oral squamous cell carcinoma (OSCC) evolution through H3K4 trimethylation (H3K4me3) and epigenetic remodeling of the chromatin landscape. Given that metabolism has also been shown to modulate cell plasticity via epigenomic modifications, we have aimed to decode the relationship between serine synthesis and cell identity in OSCC. We have shown that under serine starvation conditions, OSCC cells upregulate steady-state mRNA and protein levels of serine synthesis pathway enzymes. The latter is associated with an increase in the by-product, alpha-ketoglutarate (αKG), a co-substrate for nuclear αKG-dependent dioxygenases, which demethylate histone marker H3K27me3 and de-repress differentiation-associated genes. Furthermore, increased production of αKG is associated with downregulation of the β-catenin/CBP/MLl1 complex and CSC-associated H3K4me3. In this study, we used human OSCC cell lines CAL27 and HSC3, derived from a primary tongue tumor and a metastatic site, respectively. Each cell line was cultured in either complete medium or serine starvation medium. Serine starvation conditions led to a downregulation of H3K27me3 along with H3K4me3, promoting a switch from cancer stem cell to differentiation-enhancing chromatin landscape. The observed changes in cell plasticity were further investigated through tumorsphere formation, a surrogate assay for cancer stem cells. We found that OSCC CAL27 and HSC3 cells readily formed tumorspheres replete with stem cell markers BMI1, KRT14 and SOX2, under non-adhesive conditions when grown in complete media, supporting our findings that utilization of exogenous serine enables OSCC cells to maintain stemness. Our findings suggest that a switch from exogenous serine uptake to the endogenous serine biosynthesis promotes OSCC cell differentiation concomitant with the loss of CSC identity. Citation Format: Stacy Ann Jankowski, Nina C. Hardy, Maria A. Kukuruzinska. Exogenous serine promotes cancer stem cells in oral squamous cell carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 6049.

Oral Squamous Cell Carcinoma (OSCC) is a devastating disease associated with high morbidity, poor survival, and few therapeutic options. This malignancy is driven, in part, by epigenetic reprogramming of the chromatin landscape directed by β-catenin in complex with CREB-binding protein (CBP) and mixed lineage leukemia methyltransferase 1 (MLL1). The β-catenin/CBP/MLL1 complex promotes an open chromatin structure by enabling transcription of genes associated with cell plasticity, including cancer stem cells and cells with partial EMT (p-EMT) phenotypes. Growing evidence indicates that cell plasticity and cellular senescence are integral processes shared by cancer and aging. Given that the median age of OSCC diagnosis is 66 years, it is likely that aging promotes OSCC evolution to advanced disease. To examine the effects of aging on OSCC evolution, we adapted a syngeneic mouse model of tobacco-associated oral carcinogenesis. This model utilizes a mouse cell line, 4MOSC1, derived from 4-nitroquinoline-1 oxide- (4NQO)-induced tongue tumor which, when implanted into mouse tongues, generates tumors that recapitulate human OSCC mutanome. The growth of 4MOSC1 derived orthotopic tumors was studied in 6- and 80-week-old mice. Tumors were allowed to develop for 18 days, after which they were harvested and processed for single cell RNA sequencing, histopathology and immunofluorescence (IF) analyses. Results showed that tumors grew at a faster rate and to a greater overall size in old mice compared to their young cohorts. OSCC harvested from the old mice displayed statistically significant reduction in membranous E-cadherin concomitant with increased Cbp and H3K4me3 abundance, suggesting an age-associated upregulation of β-catenin/Cbp/Mll1 epigenetic activity. Increased expression of Bmi1, keratin 14, and podoplanin supported increased cell plasticity in OSCC from aged mice. Further, these tumors exhibited augmented cellular senescence, as judged by the disruption of lamin B1-associated nuclear membrane integrity, and by an increased cell population with markers of senescence from single cell sequencing analyses. These data suggest that aging is associated with increased β-catenin/Cbp/Mll1 epigenetic signaling, cell plasticity and cellular senescence, which collectively contribute to the evolution of OSCC. Citation Format: Emily R. Fisher, Anthony Spinella, Nina C. Hardy, Xaralabos Varelas, Manish Bais, Maria A. Kukuruzinska. Wnt/beta-catenin mediated epigenetic modifications drive age-dependent oral squamous cell carcinoma evolution. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 4743.

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