Pathways In Head And Neck Squamous Cell Carcinoma Research Articles

MLN4924 resses Head and Neck Squamous Cell Carcinoma Progression by Inactivating the mTOR Signaling Pathway via the NEDD8/CUL4/TSC2 axis.

Head and neck squamous cell carcinoma (HNSCC) is an aggressive cancer with a five-year survival rate below 50%. Standard treatments for HNSCC include surgery, radiotherapy, chemotherapy, and targeted therapies, but they still have significant limitations. Neddylation, a post-translational modification involving the attachment of NEDD8 (neural precursor cells expressed developmentally down-regulated 8) to proteins, is frequently dysregulated in HNSCC, thereby promoting tumor growth. MLN4924, also known as Pevonedistat, is a Neddylation inhibitor that has shown promise in suppressing HNSCC cell proliferation and invasion, establishing it as a potential therapeutic option. However, its precise molecular mechanism remains unclear. This study aims to investigate the mechanism of MLN4924 in HNSCC. This study examined the effects of MLN4924 on HNSCC and its associated molecular pathways. Bioinformatic analysis indicated that NEDD8, a critical component of the Neddylation pathway, is linked to poor prognosis and the mTOR (mammalian target of rapamycin) signaling pathway in HNSCC. MLN4924 significantly suppressed cell migration, invasion, and the epithelial-mesenchymal transition (EMT) pathway, and downregulated NEDD8 expression. Mechanistic studies demonstrated that MLN4924 inhibited the binding of NEDD8 to cullin4 (CUL4) and prevented the Neddylation of tuberous sclerosis complex 2 (TSC2), leading to the inactivation of the mTOR pathway. These findings were confirmed in vivo, where MLN4924 effectively inhibited tumor growth. Overall, MLN4924 disrupted Neddylation pathway and stabilized TSC2, thereby inactivating the mTOR pathway. The study provided a theoretical basis for the clinical potential of MLN4924 in improving treatment outcomes for HNSCC patients, offering a novel strategy for addressing this challenging disease.

P4HA3 promotes head and neck squamous cell carcinoma progression via the WNT/β-catenin signaling pathway

Here, we explored the role of Prolyl 4-Hydroxylase Subunit Alpha 3 (P4HA3), the most recently identified member of the prolyl-4-hydroxylase (P4H) family, in head and neck squamous cell carcinoma (HNSCC) progression. P4HA3 is upregulated during cancer progression; however, its specific role in HNSCC progression remains elusive. Thus, this study aimed to elucidate the regulatory function of P4HA3 in HNSCC development and progression and to describe the underlying mechanisms. Initially, we analyzed the correlation between the expression of P4HA3 and the WNT pathway genes and clinicopathologic features in HNSCC based on microarray data from The Cancer Genome Atlas (TCGA). Next, we used Gene Oncology (GO) functional data to describe several potentially associated pathways in HNSCC. Then, we knocked down P4HA3 in SCC15 and SCC25 cells, two classic HNSCC cell lines, and assessed the resulting changes using RT-qPCR. Furthermore, we used Western blot to evaluate the regulatory role of P4HA3 in the epithelial-to-mesenchymal transition (EMT) and the WNT/β-catenin signaling pathway. To explore the effect of P4HA3 knockdown on tumor progression, in vivo experiments were conducted using a murine model. Immunohistochemistry assays were then employed to identify proteins associated with EMT and the WNT/β-catenin signaling pathway in tumor tissues. Upregulated P4HA3 in HNSCC patient tumor tissues was positively correlated with poor prognosis. Notably, P4HA3 knockdown significantly inhibited the proliferative and invasive abilities of HNSCC. The levels of genes and proteins associated with EMT and the WNT/β-catenin signaling pathway were also markedly reduced by P4HA3 knockdown. Importantly, the in vivo experiments demonstrated that P4HA3 can promote subcutaneous tumorigenesis in nude mice and knockdown of P4HA3 induce a significant ihibitation of EMT and WNT/β-catenin pathway detected by immunohistochemistry assay in tumor tissues. In summary, we demonstrated that P4HA3 is a promising diagnostic and therapeutic biomarker for HNSCC. As an oncogene, P4HA3 increases HNSCC proliferation by inducing the EMT and activating the WNT/β-catenin signaling pathway.

The Role of the PI3K/Akt/mTOR Axis in Head and Neck Squamous Cell Carcinoma.

Head and neck squamous cell carcinoma (HNSCC) is one of the most common malignancies globally, representing a significant public health problem with a poor prognosis. The development of efficient therapeutic strategies for HNSCC prevention and treatment is urgently needed. The PI3K/AKT/mTOR (PAM) signaling pathway is a highly conserved transduction network in eukaryotic cells that promotes cell survival, growth, and cycle progression. Dysfunction in components of this pathway, such as hyperactivity of PI3K, loss of PTEN function, and gain-of-function mutations in AKT, are well-known drivers of treatment resistance and disease progression in cancer. In this review, we discuss the major mutations and dysregulations in the PAM signaling pathway in HNSCC. We highlight the results of clinical trials involving inhibitors targeting the PAM signaling pathway as a strategy for treating HNSCC. Additionally, we examine the primary mechanisms of resistance to drugs targeting the PAM pathway and potential therapeutic strategies.

LncRNA RASAL2-AS1 promotes METTL14-mediated m6A methylation in the proliferation and progression of head and neck squamous cell carcinoma

BackgroundLong non-coding RNAs (lncRNAs) are key regulators of the 6-methyladenosine (m6A) epigenetic modification, playing a role in the initiation and progression of tumors. However, the regulatory mechanisms in head and neck squamous cell carcinoma (HNSCC) remain elusive. In this study, we investigated the molecular regulatory mechanisms of the lncRNA RASAL2-AS1 in the occurrence and development of HNSCC tumors.MethodsA bioinformatics analysis was conducted to analyze the expression level of RASAL2-AS1 in HNSCC and normal tissues. RASAL2-AS1 mRNA and protein levels were detected using RT-PCR and Western blotting. Wound healing, transwell assays, flow cytometry, M6A dot blot, and RNA immunoprecipitation experiments were conducted to explore the regulatory role of the RASAL2-AS1 and downstream targets METTL14/LIS1 signaling pathway in HNSCC. Immunohistochemical examination was conducted to evaluate the expression of METTL14 and LIS1 in HNSCC and normal tissues. A tumor xenograft model of BALB/c nude mice was established to assess the impact of RASAL2-AS1 on cell proliferation and growth.ResultsRASAL2-AS1 high expression in HNSCC and cells deteriorated with survival rates of HNSCC. RASAL2-AS1 overexpression in HNSCC accelerated cell migration, colony formation, cell proliferation, cell cycle in S stage, while RASAL2-AS1 knockdown in HNSC cells inhibited cell cycle in G1 stage. After silencing METTL14, the above effects induced by overexpression of the RASAL2-AS1 were reversed. RASAL2-AS1 overexpression prompted LIS1 expression, whereas RASAL2-AS1 silencing reduced LIS1 levels in HNSCC cells, which was confirmed by immunohistological staining. Results demonstrated elevated expression of METTL14 or LIS1 in tongue cancer tissues. Overexpression of RASAL2-AS1 promoted tumor weight and tumor volume, which was counteracted by pcDNA3.1 RASAL2-AS1 plus silencing METTL14 and METTL14 and LIS1 were significantly decreased.ConclusionOur study highlights the functional importance of the LncRNA RASAL2-AS1 in HNSCC and might assist in the development of a prognostic stratification and therapeutic approach. Which regulates HNSCC with the dependence of m6a manner.

CELSR3 is a prognostic marker in HNSCC and correlates with immune cell infiltration in the tumor microenvironment

PurposeTo look at the diagnostic value of the CELSR receptor 3 (CELSR3) gene in head and neck squamous cell carcinoma (HNSCC) and its effect on tumor immune invasion, which is important for enhancing HNSCC treatment.MethodsSeveral bioinformatics tools were employed to investigate CELSR3’s putative oncogenic pathway in HNSCC, and datasets from The Tumor Genome Atlas (TCGA), Tumor Immune Estimation Resource (TIMER), Gene Expression Profile Interaction Analysis (GEPIA) and LinkedOmics were extracted and evaluated. CELSR3 has been linked to tumor immune cell infiltration, immunological checkpoints, and immune-related genes. CELSR3’s putative roles were investigated using Gene ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG), and pathway enrichment analysis. The expression level of CELSR3 in HNSCC tissues and cells was detected by RT-qPCR. The effects of CELSR3 on proliferation of HNSCC cells were detected by CCK-8 assay.ResultsCELSR3 was shown to be expressed differently in different types of cancer and normal tissues. CELSR3 gene expression was linked to pN-stage and pM-stage. Patients with high CELSR3 expression also have a well prognosis. CELSR3 expression was found to be an independent predictive factor for HNSCC in both univariate and multivariate Cox regression analyses. We discovered the functional network of CELSR3 in HNSCC using GO and KEGG analysis. CELSR3 expression levels were found to be favorably associated with immune cell infiltration levels. Furthermore, CELSR3 expression levels were significantly correlated with the expression levels of many immune molecules, such as MHC genes, immune activation genes, chemokine receptors, and chemokines. CELSR3 is highly expressed in HNSCC tissues and cells. CELSR3 overexpression significantly inhibited the proliferation of HNSCC cells. CELSR3 expression may affect the immune microenvironment and, as a result, the prognosis of HNSCC.ConclusionCELSR3 expression is elevated in HNSCC tumor tissues, and high CELSR3 expression is associated with well prognosis, which inhibited the proliferation of NHSCC cells. CELSR3 has the potential to influence tumor formation by controlling tumor-infiltrating cells in the tumor microenvironment (TME). As a result, CELSR3 may have diagnostic significance in HNSCC.

IFIT3 accelerates the progression of head and neck squamous cell carcinoma by targeting PD-L1 to activate PI3K/AKT signaling pathway

BackgroundEmerging evidence has shown interferon-induced protein with tetratricopeptide repeats 3 (IFIT3) may be predicted to be a candidate oncogene and involved in the onset and progression of cancer, but IFIT3’s potential role in cancer, particularly in head and neck squamous cell carcinoma (HNSC), is not well recognized. This study aims to reveal the role of IFIT3 in HNSC and the underlying molecular mechanism.MethodsBioinformatics analysis, immunohistochemical staining, RT-PCR, and Western blotting analysis were used to detect IFIT3 expression in HNSC. CCK-8 assays, colony formation assays, wound-healing assays, transwell assays, and sphere formation were used to explore proliferative, migratory, and invasive activities and cancer stemness of HNSC cells after IFIT3 knockdown and over-expressed. The alterations of EMT markers and PI3K/AKT pathway were detected by Western blotting. Animal studies were performed to analyze the effect of IFIT3 on tumor growth and metastasis of HNSC in vivo.ResultsIn this study, we observed that IFIT3 was highly expressed in HNSC, and its higher expression contributed to poorer survival of patients with clinical stage IV or grade 3. Function assay indicated that IFIT3 promoted malignant behaviors in vitro, as well as tumor growth and lung metastasis in vivo. Meanwhile, PD-L1 knockdown or over-expressed reversed cancer cell stemness, migration, invasion, and PI3K/AKT signaling pathway which were regulated by IFIT3.ConclusionsOur results reveal that IFIT3 promotes EMT and cancer stemness by targeting PD-L1 to activate PI3K/AKT signaling pathway in HNSC, and targeting IFIT3 may be a novel strategy for the treatment of patients with HNSC.

WNT7A promotes tumorigenesis of head and neck squamous cell carcinoma via activating FZD7/JAK1/STAT3 signaling.

Wnt signaling are critical pathway involved in organ development, tumorigenesis, and cancer progression. WNT7A, a member of the Wnt family, remains poorly understood in terms of its role and the underlying molecular mechanisms it entails in head and neck squamous cell carcinoma (HNSCC). According to the Cancer Genome Atlas (TCGA), transcriptome sequencing data of HNSCC, the expression level of WNT7A in tumors was found to be higher than in adjacent normal tissues, which was validated using Real-time RT-PCR and immunohistochemistry. Unexpectedly, overexpression of WNT7A did not activate the canonical Wnt-β-catenin pathway in HNSCC. Instead, our findings suggested that WNT7A potentially activated the FZD7/JAK1/STAT3 signaling pathway, leading to enhanced cell proliferation, self-renewal, and resistance to apoptosis. Furthermore, in a patient-derived xenograft (PDX) tumor model, high expression of WNT7A and phosphorylated STAT3 was observed, which positively correlated with tumor progression. These findings underscore the significance of WNT7A in HNSCC progression and propose the targeting of key molecules within the FZD7/JAK1/STAT3 pathway as a promising strategy for precise treatment of HNSCC.

The Role of Hedgehog Signaling Pathway in Head and Neck Squamous Cell Carcinoma.

Head and neck squamous cell carcinoma (HNSCC) is the sixth leading malignancy worldwide, with a poor prognosis and limited treatment options. Molecularly targeted therapies for HNSCC are still lacking. However, recent reports provide novel insights about many molecular alterations in HNSCC that may be useful in future therapies. Therefore, it is necessary to identify new biomarkers that may provide a better prediction of the disease and promising targets for personalized therapy. The poor response of HNSCC to therapy is attributed to a small population of tumor cells called cancer stem cells (CSCs). Growing evidence indicates that the Hedgehog (HH) signaling pathway plays a crucial role in the development and maintenance of head and neck tissues. The HH pathway is normally involved in embryogenesis, stem cell renewal, and tissue regeneration. However, abnormal activation of the HH pathway is also associated with carcinogenesis and CSC regulation. Overactivation of the HH pathway was observed in several tumors, including basal cell carcinoma, that are successfully treated with HH inhibitors. However, clinical studies about HH pathways in HNSCC are still rare. In this review, we summarize the current knowledge and recent advances regarding the HH pathway in HNSCC and discuss its possible implications for prognosis and future therapy.

P-mTOR, p-ERK and PTEN Expression in Tumor Biopsies and Organoids as Predictive Biomarkers for Patients with HPV Negative Head and Neck Cancer

BackgroundSurvival rates of head and neck squamous cell carcinoma (HNSCC) have only marginally improved in the last decades. Hence there is a need for predictive biomarkers for long-time survival that can help to guide treatment decisions and might lead to the development of new therapies. The phosphatidylinositol 3-kinase (PI3K)/AKT/mTOR signaling pathway is the most frequently altered pathway in HNSCC, genes are often mutated, amplificated and overexpressed causing aberrant signaling affecting cell growth and differentiation. Numerous genetic alterations of upstream and downstream factors have currently been clarified. However, their predictive value has yet to be established. Therefore we assess the predictive value of p-mTOR, p-ERK and PTEN expression.MethodsTissue microarrays (TMA’s) of HPV-negative patients with oropharyngeal (n = 48), hypopharyngeal (n = 16) or laryngeal (n = 13) SCC, treated with primary chemoradiation (cisplatin/carboplatin/cetuximab and radiotherapy), were histologically stained for p-mTOR, PTEN and p-ERK. Expression was correlated to overall survival (OS), disease free survival (DFS) and locoregional control (LRC). Also p-mTOR was histologically stained in a separate cohort of HNSCC organoids (n = 8) and correlated to mTOR-inhibitor everolimus response.ResultsHigh p-mTOR expression correlated significantly with worse OS in multivariate analysis in the whole patient cohort [Hazar Ratio (HR) 1.06, 95%CI 1.01–1.11, p = 0.03] and in the cisplatin/carboplatin group with both worse OS (HR 1.09, 95%CI 1.02–1.16, p = 0.02) and DFS (HR 1.06, 95%CI 1.00–1.12, p = 0,04). p-ERK expression correlated significantly with DFS in univariate analysis in the whole patient cohort (HR 1.03, 95%CI 1.00–1.05, p = 0.04) and cisplatin/carboplatin group (HR 1.03, 95%CI 1.00–1.07, p = 0.04). PTEN-expression did not correlate with OS/DFS/LRC. Better organoid response to everolimus correlated significantly to higher p-mTOR expression (Rs = − 0.731, p = 0.04).ConclusionsHigh p-mTOR expression predicts and high p-ERK expression tends to predict worse treatment outcome in HPV negative HNSCC patients treated with chemoradiation, providing additional evidence that these markers are candidate prognostic biomarkers for survival in this patient population. Also this study shows that the use of HNSCC organoids for biomarker research has potential. The role of PTEN expression as prognostic biomarker remains unclear, as consistent evidence on its prognostic and predictive value is lacking.

Abstract 4951: Proteasome inhibitor carfilzomib induces apoptosis via downregulation of AKT pathway in HPV negative HNSCC

Abstract Background Head and neck squamous cell carcinoma (HNSCC) presents in heterogenous sites, varied etiology and risk factors, and has diverse genetic background which confer dissimilar response to treatments, including chemotherapy. Here we tested the hypothesis that investigation of HPV presence in a HNSCC cell line would identify pathway contributing to oncogenesis of other HNSCC. We sought to identify pathways that differentially affect HNSCC according to the genetic background, therefore identifying targets and suitable candidate for treatment. We used genetic and pharmacological approaches to inhibit AKT pathway in HNSCC cell lines. Materials & methods Chemical genetic library screening against 6 HNSCC cell lines were performed. The high throughput combinatorial small molecule screening for cell viability and apoptosis was carried out. Candidate agents were validated in both CAL-27 (HPV-negative) and UM-SCC-47 (HPV-positive) cell lines. Identified pathways and representative targeted agents were validated in vitro. Expression of related protein were confirmed by western blot assay. Results EGFR, PIK3CA, mTOR, CDK1, HSP90AB1, HDAC1, and PSMD1 pathways were identified to be susceptible pathways in HNSCC using high throughput chemical genetic library screening. In vitro validation of carfilzomib showed IC50 of 7.07nM in CAL-27 cells and IC50 of 26.3nM in UM-SCC-47 cells. Knockdown of PSMD1 transcripts by transfected siRNAs reduced CAL-27 cells viability. Carfilzomib regulates the AKT pathway by decreasing t-AKT, p-AKT, p-ERK, p-S6 proteins and increasing p21 in CAL-27. Conclusion Inhibition of AKT pathway treatment with proteasome inhibitor carfilzomib is effective in HPV-negative HNSCC cells. Taken together, our findings reveal carfilzomib induced the apoptotic pathway and suggest that carfilzomib, the representative of a new class of chemotherapeutic drug, may be utilized for overcoming cisplatin-resistance in human SCC. Citation Format: Woo-Jin Jeong, Jiyeong Kim, Hye-Yeon Lee, Zhiyong Wang, Walt Amornphimoltham, Silvio Gutkind. Proteasome inhibitor carfilzomib induces apoptosis via downregulation of AKT pathway in HPV negative HNSCC. [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 4951.

Abstract 384: Secondhand smoke exposure alters cisplatin-induced cell death and alters cell death and survival pathway in HNSCC

Abstract BACKGROUND: Secondhand smoke (SHS) contains more than 7000 chemicals, 70of which are carcinogens and it is estimated that every one in four Americans areinvoluntarily exposed to SHS. Carcinogenesis induced by SHS exposure isunclear, though SHS exposure is associated with various cancers including headand neck squamous cell carcinoma (HNSCC). Recently, it has been shown thatSHS exposure is a significant independent predictor of HNSCC recurrence. Wehave shown that in vitro exposure to SHS increases HNSCC cisplatin resistance. AIMS: Our study examines the effect of SHS smoke exposure on cisplatin-induced HNSCC cancer cell death and the possible mechanisms behind thisobservation. METHODS: Sidestream smoke (SS), the main component of SHS,was extracted as previously described. Authenticated HNSCC cell lines (UM-SCC1 & WSU-HN6) were exposed to SS extract for 48 hours at doses mimickingthe nicotine levels observed in the saliva of passive smokers. Then, cancer cellswere treated with cisplatin in the presence or absence of SS extract. Cisplatin-induced cancer cell death was measured using Incucyte® S3 Live-Cell imaginganalysis. Transcriptomic analysis following RNA-sequencing was performedusing Ingenuity Pathway Analysis. Data were subjected to ANOVA. RESULTS: Our live-cell imaging system revealed that HNSCC cells exposed to SS extractsignificantly decreased cisplatin-induced cancer cell death at various time points compared with cells treated with cisplatin alone in both cell lines. The HNSCCcells treated only with SS extract did not show any change in cell death at 0, 12,24, 48, 72, and 96 hours. RNA-sequencing data revealed that cell death andsurvival pathways are the most changed pathways in the SS extract-treatedgroup compared with unexposed HNSCC cells. CONCLUSIONS: Our studydocuments for the first time that even short-term exposure to SHS allows cells toevade cisplatin-induced cell death and can lead to head and neck cancercisplatin resistance. At the molecular level, SHS exposure alters genes that areassociated with cell death and survival. Further, HNSCC patients-basedobservations and mechanistic studies are warranted. FUNDING: This work was partially supported by the National Cancer Institute of the NationalInstitutes of Health (R33CA202898, R01CA242168, and P30CA225520), theTSET Health Promotion Cancer Center, and the Oklahoma Center forAdvancement of Science and Technology (HR16-007). Dr. Queimado holds aPresbyterian Health Foundation Endowed Chair in Otorhinolaryngology Position. Citation Format: Balaji Sadhasivam, Jimmy Manyanga, Vengatesh Ganapathy, Célia Bouharati, David Rubenstein, Jonathan Wren, Pawan Acharya, Daniel Zhao, Lurdes Queimado. Secondhand smoke exposure alters cisplatin-induced cell death and alters cell death and survival pathway in HNSCC [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 384.

FAM3D as a Prognostic Indicator of Head and Neck Squamous Cell Carcinoma Is Associated with Immune Infiltration.

Globally, head and neck squamous cell carcinoma (HNSCC) is a common malignant tumor with high morbidity and mortality. Hence, it is important to find effective biomarkers for the diagnosis and prediction of the prognosis of patients with HNSCC. FAM3D had been proven to be vital in other cancers. However, its predictive and therapeutic value in HNSCC is unclear. Therefore, it is valuable to explore the association between the expression level of FAM3D and its impacts on the prognosis and tumor microenvironment in HNSCC. The Cancer Genome Atlas (TCGA) dataset, Genotype-Tissue Expression (GTEx) dataset, the Clinical Proteomic Tumor Analysis Consortium (CPTAC) dataset, and The Human Protein Atlas (THPA) website were used to assess HNSCC expressions in tumor and nontumor tissues. Then, we further conducted immunohistochemistry experiment as internal cohort to validate the same results. The Cox regression analysis, Kaplan-Meier analysis, and nomograms were performed to find the predictive prognostic value of FAM3D in HNSCC patients and its relationship with the clinicopathological features in HNSCC. The Gene Expression Omnibus (GEO) dataset was utilized to externally verify the prognosis value of FAM3D in HNSCC. Gene Set Enrichment Analysis (GESA) was applied to search the molecular and biological functions of FAM3D. The association between FAM3D and immune cell infiltration was investigated with the Tumor Immune Estimating Resource, version 2 (TIMER2). The relationships between FAM3D expression and tumor microenvironment (TME) scores, immune checkpoints, and antitumor compound half-maximal inhibitory concentration predictions were also explored. In different datasets, FAM3D mRNA and protein levels were all significantly lower in HNSCC tissues than in normal tissues, and they were strongly inversely associated with tumor grade, stage, lymph node metastasis, and T stage. Patients with high-FAM3D-expression displayed better prognosis than those with low-FAM3D-expression. FAM3D was also determined to be a suitable biomarker for predicting the prognosis of patients with HNSCC. This was externally validated in the GEO dataset. As for gene and protein level, the functional and pathway research results of FAM3D indicated that it was enriched in alteration of immune-related pathways in HNSCC. The low-expression group had higher stromal and ESTIMATE scores by convention than the high-expression group. FAM3D expression were found to be positively correlated with immune infiltrating cells, such as cancer-associated fibroblasts, myeloid-derived suppressor cells, macrophage cells, T cell CD8+ cells, regulatory T cells, and T cell follicular helper cells. FAM3D's relationships with immune checkpoints and sensitivity to antitumor drugs were also investigated. Our study explored the impact of FAM3D as a favorable prognostic marker for HNSCC on the tumor immune microenvironment from multiple perspectives. The results may provide new insights into HNSCC-targeted immunotherapy.

ADAMTS12, a novel prognostic predictor, promotes cell proliferation, migration, and invasion in head and neck squamous cell carcinoma.

The prognostic significance and potential carcinogenic mechanism of ADAM metallopeptidase with thrombospondin type 1 motif 12 (ADAMTS12) in head and neck squamous cell carcinoma (HNSC) remain unclear. Immunohistochemistry was used to analyze the correlation between ADAMTS12 protein expression and clinicopathological factors in tumor samples from 195 patients with HNSC. Based on clinicopathological data of patients, Cox regression and Kaplan-Meier analysis were used to identify the prognostic significance of the ADAMTS12 expression. The carcinogenicity of the ADAMTS12 in HNSC cells was analyzed by CCK-8 assay, the wound-healing assay, and transwell assays after transfection of ADAMTS12 overexpression or knock-down vector. The expression of ADAMTS12 was up-regulated in HNSC compared with normal tissue, related to pathology grade and lymph node metastasis of patients with HNSC, which was an independent prognostic factor. ADAMTS12 overexpression facilitated cell viability, invasion, and migration of HNSC cells, while ADAMTS12 knock-down had inverse results. Moreover, enrichment analysis, ADAMTS12 overexpression assay, and ADAMTS12 knock-down assay confirmed that ADAMTS12 mediated the activation of P13K/Akt pathway in HNSC. Our studies indicated that ADAMTS12 was a novel prognostic biomarker and potentially therapeutic target in HNSC.

Integrating Genetic Alterations and the Hippo Pathway in Head and Neck Squamous Cell Carcinoma for Future Precision Medicine.

Genetic alterations and dysregulation of signaling pathways are indispensable for the initiation and progression of cancer. Understanding the genetic, molecular, and signaling diversities in cancer patients has driven a dynamic change in cancer therapy. Patients can select a suitable molecularly targeted therapy or immune checkpoint inhibitor based on the driver gene alterations determined by sequencing of cancer tissue. This “precision medicine” approach requires detailed elucidation of the mechanisms connecting genetic alterations of driver genes and aberrant downstream signaling pathways. The regulatory mechanisms of the Hippo pathway and Yes-associated protein/transcriptional co-activator with PDZ binding motif (YAP/TAZ) that have central roles in cancer cell proliferation are not fully understood, reflecting their recent discovery. Nevertheless, emerging evidence has shown that various genetic alterations dysregulate the Hippo pathway and hyperactivate YAP/TAZ in cancers, including head and neck squamous cell carcinoma (HNSCC). Here, we summarize the latest evidence linking genetic alterations and the Hippo pathway in HNSCC, with the aim of contributing to the continued development of precision medicine.

Decursin inhibits tumor progression in head and neck squamous cell carcinoma by downregulating CXCR7 expressionin vitro

CXC chemokine receptor 7 (CXCR7) is frequently overexpressed in cancer and plays a significant role in tumor growth and metastasis. Consequently, inhibition of CXCR7 is important for treatment strategies. However, little is known concerning the biological role of CXCR7 and its underlying mechanisms in head and neck squamous cell carcinoma (HNSCC). The present study investigated the role of CXCR7 in HNSCC, as well as the effects of decursin, a pyranocoumarin compound isolated from Angelica gigas Nakai, on CXCR7 and its downstream signaling. Expression levels of CXCR7 in HNSCC cells were examined using flow cytometry, reverse transcriptase PCR, western blot analysis, and immunofluorescence. The effects of CXCR7 on cell proliferation, migration, and invasion were studied using CCK-8, gap closure, and transwell assays. The results revealed that decursin significantly reduced CXCR7 expression and inhibited cell proliferation, migration, and invasion of human HNSCC cell lines. In addition, decursin induced G0/G1 cell cycle arrest in CXCR7-overexpressing cells and decreased the levels of cyclin A, cyclin E, and CDK2. Furthermore, CXCR7 promoted cancer progression via the STAT3/c-Myc pathway in HNSCC; suppression of CXCR7 with decursin prevented this effect. These results suggest that CXCR7 promotes cancer progression through the STAT3/c-Myc pathway and that the natural compound decursin targets CXCR7 and may be valuable in the treatment of HNSCC.

Identification of key genes for HNSCC from public databases using bioinformatics analysis

BackgroundThe cause and underlying molecular mechanisms of head and neck squamous cell carcinoma (HNSCC) are unclear. Our study aims to identify the key genes associated with HNSCC and reveal potential biomarkers.MethodsIn this study, the expression profile dataset GSE83519 of the Gene Expression Omnibus database and the RNA sequencing dataset of HNSCC of The Cancer Genome Atlas were included for analysis. Sixteen differentially expressed genes were screened from these two datasets using R software. Gene Expression Profiling Interactive Analysis 2 (GEPIA2) was then adopted for survival analysis, and finally, three key genes related to the overall survival of HNSCC patients were identified. Furthermore, we verified these three genes using the Oncomine database and from real-time PCR and immunohistochemistry results from HNSCC tissues.ResultsThe expression data of 44 samples from GSE83519 and 545 samples from TCGA-HNSC were collected. Using bioinformatics, the two databases were integrated, and 16 DEGs were screened out. Gene Ontology (GO) enrichment analysis showed that the biological functions of DEGs focused primarily on the apical plasma membrane and regulation of anoikis. Kyoto Encyclopedia of Genes and Genomes (KEGG) signalling pathway analysis showed that these DEGs were mainly involved in drug metabolism-cytochrome P450 and serotonergic synapses. Survival analysis identified three key genes, CEACAM5, CEACAM6 and CLCA4, that were closely related to HNSCC prognosis. The Oncomine database, qRT–PCR and IHC verified that all 3 key genes were downregulated in most HNSCC tissues compared to adjacent normal tissues.ConclusionsThis study indicates that integrated bioinformatics analyses play an important role in screening for differentially expressed genes and pathways in HNSCC, helping us better understand the biomarkers and molecular mechanism of HNSCC.

FAM83A promotes proliferation and metastasis via Wnt/\u03b2-catenin signaling in head neck squamous cell carcinoma

This research aimed to investigate the expression and function of FAM83A in the proliferation and metastasis in head and neck squamous cell carcinoma (HNSCC). FAM83A mRNA and protein expressions in HNSCC were detected in primary HNSCC samples and cell lines. The associations between FAM83A expression and clinicopathologic variables were evaluated through tissue microarrays. Besides, FAM83A knockdown and overexpression cell lines were constructed to assess cell growth and metastasis in vitro and the relationship between FAM83A and epithelial-mesenchymal transition (EMT). Furthermore, two models of xenograft tumors in nude mice were used to assess the tumorigenicity and metastasis ability of FAM83A in vivo. In the present study, overexpression of FAM83A in HNSCC samples was significantly associated with tumor size, lymph node status and clinical tumor stages. Mechanically, FAM83A could promote HNSCC cell growth and metastasis by inducing EMT via activating Wnt/β-catenin signaling pathway. Rescue experiment demonstrated the inhibition of β-catenin could counteract the function of FAM83A. Also, the FAM83A knockdown could suppress tumor growth and distant metastasis in the xenograft animal models of HNSCC. In conclusion, this study identifies FAM83A as an oncogene of HNSCC. This study provides new insights into the molecular pathways that contribute to EMT in HNSCC. We revealed a previously unknown FAM83A-Wnt–β-catenin signaling axis involved in the EMT of HNSCC. There may be a potential bi-directional signaling loop between FAM83A and Wnt/β-catenin signaling pathway in HNSCC.

Lactate secreted by PKM2 upregulation promotes Galectin-9-mediated immunosuppression via inhibiting NF-\u03baB pathway in HNSCC

Pyruvate kinase M2 as a key rate-limiting enzyme in glycolysis, it plays a critical role in metabolic reprogramming and carcinogenesis. However, whether PKM2 can promote immunosuppressive microenvironment formation remains unknown in head and neck squamous cell carcinoma (HNSCC). PKM2 expression was detected using immunohistochemical staining. The biological functions of PKM2 were investigated in vitro and in vivo. Lactate production and the expression of Galectin-9, a critical immunosuppression molecule, were detected after PKM2 knockdown and overexpression in HNSCC cells. The mechanism of lactate regulating Galectin-9 expression through NF-κB signaling was explored in vitro. Overexpression of PKM2 correlates with poor prognosis in HNSCC patients. Silencing PKM2 markedly inhibits proliferation and metastasis capacity in vivo and in vitro, and vice versa. The glycolysis and glycolytic capacity are significantly decreased after PKM2 silencing. Lactate secretion induced by PKM2 significantly promotes migration and invasion capacity. Furthermore, a positive correlation between PKM2 and Galectin-9 expression is observed in HNSCC tissues. The induction of Galectin-9 expression by PKM2 can be affected by a lactate transporter inhibitor. Mechanically, lactate impeded the suppressive transcriptional complex formation of NF-κB and histone deacetylase 3 (HDAC3), which released the transcription of Galectin-9 mediated by NF-κB signaling. Our findings demonstrate that lactate produced by PKM2 upregulation promotes tumor progression and Galectin-9-mediated immunosuppression via NF-κB signaling inhibition in HNSCC, which bridges metabolism and immunosuppression. The novel PKM2-lactate-Galectin-9 axis might be a potential therapeutic target in HNSCC.

Abstract PO-066: Relationship between KEAP1/NFE2L2 pathway activation and radiation resistance in oral cavity cancer

Abstract Radiotherapy (RT) is one of the primary treatment modalities for head and neck squamous cell carcinoma (HNSCC) either in the definitive or the adjuvant setting. Unfortunately, despite therapeutic and technological advances in radiotherapy, some patients will have persistence tumor or develop locoregional failure, resulting in significant morbidity and mortality. Presently, there is no biomarker that can reliably identify HNSCC that is resistant to RT. Recent genomic sequencing studies show that mutations in the KEAP1/NFE2L2 pathway exist in HNSCC. However, very little data is available with regards to the relationship between RT resistance and activating mutation or pathway hyperactivation of this pathway in HNSCC. Moreover, the mechanism by which dysregulation of this pathway can lead to RT resistance in HNSCC is poorly understood. Therefore, the project is aimed to study the relationship between KEAP1/NFE2L2 mutations and patient outcomes in HNSCC. To address this aim, DNA sequencing strategies and tissue microarray (TMA) are used to identify mutations and/or a hyperactivation protein signature of the KEAP1/NFE2L2 oxidative stress pathway in an oral cavity cancer patient cohorts (N = 69[MOU1]) treated with surgery and adjuvant RT. In addition, identified mutations are introduced into HNSCC cell lines and mouse xenografts to study the effect of these mutations on radiation response and to identify the mechanism by which they mediate radiation resistance. Here we find that 6% of oral cavity cancer patients harbor mutation in this pathway, primarily in a hotspot in the NFE2L2 gene. NFE2L2 mutation is associated with 100% in-field recurrence after radiotherapy. Strong co-expression of two downstream targets of this pathway (HO1 and NQO1) in tumor cells on the TMA strongly correlates with NFE2L2 mutation and in-field recurrence after RT. Finally, overexpression of the mutated NFE2L2 gene in HNC cell lines lead to increased ROS and radioresistance. Our findings suggest oral cavity cancers harboring activating mutation or a hyperactivation protein signature of the KEAP1/NFE2L2 pathway are more radioresistant. Work is ongoing to validate these findings in other types of HNSCC and to determine how to enhance radiation effectiveness in mutated tumors. Citation Format: Li Guan, Hongbin Cao, Angela Hui, Shirley Kwok, Vignesh Viswanathan, Dhanya Nambiar, Rie V. Eyben, Brittany Holmes, Christina Kong, Maximilian Diehn, Quynh-Thu Le. Relationship between KEAP1/NFE2L2 pathway activation and radiation resistance in oral cavity cancer [abstract]. In: Proceedings of the AACR Virtual Special Conference on Radiation Science and Medicine; 2021 Mar 2-3. Philadelphia (PA): AACR; Clin Cancer Res 2021;27(8_Suppl):Abstract nr PO-066.

Immune Checkpoints Pathways in Head and Neck Squamous Cell Carcinoma.

Simple SummaryDuring the last decades, scientific advances in immuno-oncology and a better understanding of tumors’ immune profile led to the development of novel immunotherapeutic strategies, especially immune checkpoint inhibitors. The blockade of PD-1 by monoclonal antibodies (mAbs) is the only immunotherapy based on immune checkpoint pathways approved for head and neck squamous cell carcinoma. As only a small fraction of patients perceives clinical benefit, understanding the molecular mechanisms and signaling pathways activated by the immune checkpoints and other tumor intrinsic features that modulate the immune infiltrate is crucial to better select patients for immunotherapy treatment and to develop novel therapeutic strategies. We here review the immune escape mechanisms of head and neck tumors, with a particular focus on the immune checkpoints, their role as therapeutic targets, and the predictive biomarkers of response to anti-PD-1/PD-L1 therapy. We also summarize the ongoing clinical trials testing several combinations of immune checkpoint inhibitors with other therapeutic approaches to improve patient outcomes.Head and neck squamous cell carcinoma (HNSCC) is a heterogeneous group of tumors usually diagnosed at an advanced stage and characterized by a poor prognosis. The main risk factors associated with its development include tobacco and alcohol consumption and Human Papillomavirus (HPV) infections. The immune system has a significant role in the oncogenesis and evolution of this cancer type. Notably, the immunosuppressive tumor microenvironment triggers immune escape through several mechanisms. The improved understanding of the antitumor immune response in solid tumors and the role of the immune checkpoint molecules and other immune regulators have led to the development of novel therapeutic strategies that revolutionized the clinical management of HNSCC. However, the limited overall response rate to immunotherapy urges identifying predictive biomarkers of response and resistance to treatment. Here, we review the role of the immune system and immune checkpoint pathways in HNSCC, the most relevant clinical findings linked to immunotherapeutic strategies and predictive biomarkers of response and future treatment perspectives.