EP4-induced mitochondrial localization and cell migration mediated by CALML6 in human oral squamous cell carcinoma.

Lymph node metastasis, primarily caused by the migration of oral squamous cell carcinoma (OSCC) cells, stands as a crucial prognostic marker. We have previously demonstrated that EP4, a subtype of the prostaglandin E2 (PGE2) receptor, orchestrates OSCC cell migration via Ca2+ signaling. The exact mechanisms by which EP4 influences cell migration through Ca2+ signaling, however, is unclear. Our study aims to clarify how EP4 controls OSCC cell migration through this pathway. We find that activating EP4 with an agonist (ONO-AE1-473) increased intracellular Ca2+ levels and the migration of human oral cancer cells (HSC-3), but not human gingival fibroblasts (HGnF). Further RNA sequencing linked EP4 to calmodulin-like protein 6 (CALML6), whose role remains undefined in OSCC. Through protein-protein interaction network analysis, a strong connection is identified between CALML6 and calcium/calmodulin-dependent protein kinase kinase 2 (CaMKK2), with EP4 activation also boosting mitochondrial function. Overexpressing EP4 in HSC-3 cells increases experimental lung metastasis in mice, whereas inhibiting CaMKK2 with STO-609 markedly lowers these metastases. This positions CaMKK2 as a potential new target for treating OSCC metastasis. Our findings highlight CALML6 as a pivotal regulator in EP4-driven mitochondrial respiration, affecting cell migration and metastasis via the CaMKK2 pathway.

A peptide binding to the tetraspanin CD9 reduces cancer metastasis.

As an organizer of multi-molecular membrane complexes, the tetraspanin CD9 has been implicated in a number of biological processes, including cancer metastasis, and is a candidate therapeutic target. Here, we evaluated the suppressive effects of an eight-mer CD9-binding peptide (CD9-BP) on cancer cell metastasis and its mechanisms of action. CD9-BP impaired CD9-related functions by adversely affecting the formation of tetraspanin webs-networks composed of CD9 and its partner proteins. The anti-cancer metastasis effect of CD9-BP was evidenced by the in vitro inhibition of cancer cell migration and invasion as well as exosome secretion and uptake, which are essential processes during metastasis. Finally, using a mouse model, we showed that CD9-BP reduced lung metastasis in vivo. These findings provide insight into the mechanism by which CD9-BP inhibits CD9-dependent functions and highlight its potential application as an alternative therapeutic nano-biomaterial for metastatic cancers.

The Expression of SIRT6 Is Associated With Treatment Outcome in Elder Patients With Oral Cancer.

BACKGROUND/AIM: SIRT6 is one of seven human sirtuin genes and is known to act as an onco-suppressor gene in colorectal and ovarian cancers, although it is up-regulated in other cancers. Thus, SIRT6 is considered performing both tumor-suppressing and promoting roles. However, the association of SIRT6 with oral squamous cell carcinoma (OSCC) and its role in OSCC pathogenesis is currently unclear. This study aimed to investigate the expression of SIRT6 in patients with OSCC and its potential as a biomarker for early detection and prognosis prediction. MATERIALS AND METHODS: Immunohistochemistry, quantitative real-time RT-PCR, and microarray analyses were performed to determine SIRT6 expression and its association with clinicopathological features in OSCC using clinical specimens. RESULTS: SIRT6 mRNA and protein expression levels were higher in OSCC tissues than in noncancerous tissues (p<0.05). SIRT6 expression was predominant in patients aged ≥65 years and significantly correlated with shorter overall survival. In the microarray analysis, some SIRT6-associated genes, such as ANXA2, were up-regulated in OSCC. CONCLUSION: SIRT6 plays a role in tumor homeostasis, leading to a poor prognosis in OSCC. SIRT6 may represent a novel target not only for treatment, but also as a prognostic marker in OSCC.

Clinical investigation of patients with jaw deformity with comorbidities.

BACKGROUND: With improvements in the safety and stability of surgeries, the number of orthognathic surgeries is increasing. Most patients who undergo orthognathic surgeries are younger, and the number of orthognathic surgeries for patients with comorbidities is also increasing. We report a survey and clinical investigation of patients with comorbidities who underwent orthognathic surgeries at our department to improve the safety of orthognathic surgery. RESULTS: The participants included 296 men and 712 women, with a mean age of 28 years (13-19 years, n=144; 20-29 years, n=483; 30-39 years, n=236; 40-49 years, n=102; 50-59 years, n=39; ≥60 years, n=4). In total, 347 patients underwent one-stage Le Fort type I osteotomy and sagittal split ramus osteotomy (SSRO), 243 underwent SSRO, 287 underwent plate removal, 126 underwent genioplasty and plate removal, and five underwent other surgeries. In total, 529 patients had comorbidities (52%), including allergic diseases (n=220, 33%), respiratory diseases (n=107, 16%), neurologic and psychiatric diseases (n=69, 10%), gynecologic diseases (n=28, 4%), hematologic diseases (n=27, 4%), cardiovascular diseases (n=24, 4%), digestive diseases (n=22, 3%), metabolic and endocrine diseases (n=18, 3%), spinal diseases (n=11, 2%), ophthalmologic diseases (n=11, 2%), renal and urological diseases (n=9, 1%), and other diseases (n=117, 18%). Among the patients with comorbidities, 11 with hemorrhagic diatheses (hemophilia and von Willebrand disease), arrhythmia (atrioventricular block), psychiatric disease (adjustment disorder), and metabolic disease (diabetes) required cautious perioperative management. The patient with hemophilia was managed with regular low-dose recombinant factor VIII replacement therapy, and the patient with type I diabetes mellitus was administered continuous insulin infusion and sliding-scale insulin therapy; both patients had an uneventful course. CONCLUSIONS: The study findings suggest that with the increase in orthognathic surgeries, oral and maxillofacial surgeons should adequately manage cases requiring cautious perioperative control and highlight the importance of preoperative screening. Despite the well-established safety and postoperative stability of orthognathic surgeries, oral surgeons should adopt appropriate additional preventive measures for patients with comorbidities.

[Exosome, a Nano-Sized Carrier, and the Brain-Gut Correlation].

Exosomes, small bilayer vesicles secreted by all cells, are recognized as a novel communication tool among distant organs. They can cross the blood-brain barrier and be taken up by specific brain cells, altering the brain microenvironment and possibly playing a role in pathological conditions such as brain metastasis and neurodegenerative diseases. Intestinal bacteria also release small membrane vesicles called "bacterial extracellular vesicles" (BEVs), which can invade blood vessels and affect distant tissues in a manner similar to exosomes, suggesting that BEVs also play a role in disease progression in the brain-gut axis. Here, we will discuss the latest findings of the relationship between exosomes, including BEVs, and multiple brain pathologies.

Hypoxia and CD11b+ Cell Influx Are Strongly Associated With Lymph Node Metastasis of Oral Cancer.

BACKGROUND/AIM: Treatment failure in oral cancer is mainly caused by uncontrolled cervical lymph node (LN) metastasis. We previously reported that CD11b+ cells are recruited into tumor hypoxic areas following radiation, leading to re-vascularization and relapse. Since lymphatic vessel formation has similarities with vascular formation, we examined whether surgery induces hypoxia and stimulates lymphangiogenesis. MATERIALS AND METHODS: The recruitment of CD11b+ cells and the formation of lymphatic vessels were examined using orthotopic tongue cancer mouse models with glossectomy. RESULTS: Surgery on OSC-19 tumor induced LN metastases and hypoxia, followed by CD11b+ cell influx. These phenomena were not observed in the no tumor or SAT tumor models. Stimulation of lymphangiogenesis was observed in the CD11b+ cell influx area, as the tumor grew. The localization of CD11b+ cells was changed from the lymph nodules to the medullary sinuses. CONCLUSION: Surgery-induced hypoxia in oral tumors leads to CD11b+ cell infiltration, lymphangiogenesis, and LN metastasis.

Extracellular Vesicle and Particle Biomarkers Define Multiple Human Cancers.

There is an unmet clinical need for improved tissue and liquid biopsy tools for cancer detection. We investigated the proteomic profile of extracellular vesicles and particles (EVPs) in 426 human samples from tissue explants (TEs), plasma, and other bodily fluids. Among traditional exosome markers, CD9, HSPA8, ALIX, and HSP90AB1 represent pan-EVP markers, while ACTB, MSN, and RAP1B are novel pan-EVP markers. To confirm that EVPs are ideal diagnostic tools, we analyzed proteomes of TE- (n = 151) and plasma-derived (n = 120) EVPs. Comparison of TE EVPs identified proteins (e.g., VCAN, TNC, and THBS2) that distinguish tumors from normal tissues with 90% sensitivity/94% specificity. Machine-learning classification of plasma-derived EVP cargo, including immunoglobulins, revealed 95% sensitivity/90% specificity in detecting cancer. Finally, we defined a panel of tumor-type-specific EVP proteins in TEs and plasma, which can classify tumors of unknown primary origin. Thus, EVP proteins can serve as reliable biomarkers for cancer detection and determining cancer type.

M2-polarized macrophages contribute to neovasculogenesis, leading to relapse of oral cancer following radiation.

Despite the fact that radiation is one of the standard therapies in the treatment of patients with oral cancer, tumours can recur even in the early stages of the disease, negatively impacting prognosis and quality of life. We previously found that CD11b(+) bone marrow-derived cells (BMDCs) were recruited into human glioblastoma multiforme (GBM), leading to re-organization of the vasculature and tumour regrowth. However, it is not yet known how these cells contribute to tumour vascularization. In the present study, we investigated the role of infiltrating CD11b(+) myeloid cells in the vascularization and recurrence of oral squamous cell carcinoma (OSCC). In a xenograft mouse model, local irradiation caused vascular damage and hypoxia in the tumour and increased infiltration of CD11b(+) myeloid cells. These infiltrating cells showed characteristics of M2 macrophages (M2Mφs) and are associated with the promotion of vascularization. M2Mφs promoted tumour progression in recurrence after irradiation compared to non-irradiated tumours. In addition, we found that CD11b(+) myeloid cells, as well as CD206(+) M2Mφs, are increased during recurrence after radiotherapy in human OSCC specimens. Our findings may lead to the development of potential clinical biomarkers or treatment targets in irradiated OSCC patients.

Establishment of successively transplantable rabbit VX2 cancer cells that express enhanced green fluorescent protein.

Morphological detection of cancer cells in the rabbit VX2 allograft transplantation model is often difficult in a certain region such as serosal cavity where reactive mesothelial cells mimic cancer cells and both cells share common markers such as cytokeratins. Therefore, tagging VX2 cells with a specific and sensitive marker that easily distinguishes them from other cells would be advantageous. Thus, we tried to establish a successively transplantable, enhanced green fluorescent protein (EGFP)-expressing VX2 model. Cancer cells obtained from a conventional VX2-bearing rabbit were cultured in vitro and transfected with an EGFP-encoding vector, and then successively transplanted in Healthy Japanese White rabbits (HJWRs) (n = 8). Besides, conventional VX2 cells were transplanted in other HJWRs (n = 8). Clinicopathological comparison analyses were performed between the two groups. The success rate of transplantation was 100% for both groups. The sensitivity and specificity of EGFP for immunohistochemical detection of VX2 cells were 84.3 and 100%, respectively. No significant differences in cancer cell morphology, tumor size (P = 0.742), Ki-67 labeling index (P = 0.878), or survival rate (P = 0.592) were observed between the two. VX2 cells can be genetically altered, visualized by EGFP, and successively transplanted without significant alteration of morphological and biological properties compared to those of the conventional model.