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Radiotherapy plays an essential role in the treatment of head and neck squamous cell carcinoma (HNSCC), yet radioresistance remains a major barrier to therapeutic efficacy. A better understanding of the predominant pathways determining radiotherapy response could help develop mechanism-informed therapies to improve cancer management. Here we report that radioresistant HNSCC cells exhibit increased tumor aggressiveness. Using unbiased proteome profiler antibody arrays, we identify that upregulation of c-Met phosphorylation is one of the critical mechanisms for radioresistance in HNSCC cells. We further uncover that radioresistance-associated HNSCC aggressiveness is effectively exacerbated by c-Met but is suppressed by its genetic knockdown and pharmacologic inactivation. Mechanistically, the resulting upregulation of c-Met promotes elevated expression of plexin domain containing 2 (PLXDC2) through activating ERK1/2-ELK1 signaling, which in turn modulates cancer cell plasticity by epithelial–mesenchymal transition (EMT) induction and enrichment of the cancer stem cell (CSC) subpopulation, leading to resistance of HNSCC cells to radiotherapy. Depletion of PLXDC2 overcomes c-Met–mediated radioresistance through reversing the EMT progress and blunting the self-renewal capacity of CSCs. Therapeutically, the addition of SU11274, a selective and potent c-Met inhibitor, to radiation induces tumor shrinkage and limits tumor metastasis to lymph nodes in an orthotopic mouse model. Collectively, these significant findings not only demonstrate a novel mechanism underpinning radioresistance-associated aggressiveness but also provide a possible therapeutic strategy to target radioresistance in patients with HNSCC. Significance: This work provides novel insights into c-Met-PLXDC2 signaling in radioresistance-associated aggressiveness and suggests a new mechanism-informed therapeutic strategy to overcome failure of radiotherapy in patients with HNSCC.

Coronary microvascular dysfunction contributes to the development of left ventricular (LV) diastolic dysfunction in heart failure with preserved ejection fraction (HFpEF), a prevalent symptom that has limited targeted treatment options. Selective pharmacological inhibitors of phosphodiesterase 9A (PDE9A), such as PF04447943, have emerged as a potential therapeutic approach for the treatment of cardiometabolic diseases, in part, owing to their antioxidant effects. We hypothesized that PDE9A inhibition has beneficial effects on the coronary microvasculature, thereby improving LV diastolic function in HFpEF. In order to study this, the Obese diabetic Zucker fatty/spontaneously hypertensive heart failure F1 hybrid (ZSF1) rat, a model of human HFpEF, was employed in which it was found that progressively developing coronary microvascular rarefaction, as quantified by an unbiased, three-dimensional reconstruction of the coronary microvascular network (Vesselucida360), as well as vasodilator coronary arteriole dysfunction are associated with the development of LV diastolic dysfunction, which were improved by the treatment with the selective PDE9A inhibitor, PF04447943 (3 mg/kg/day, oral gavage for 2 weeks). These coronary microvascular changes were accompanied by an increase in cardiac 3-nitrotyrosine, a marker of nitrosative stress, which was also mitigated by PF04447943 treatment. Furthermore, mass spectrometry-based proteomics and pathway analyses identified peroxiredoxin-5 (PRDX5) as one of the predominantly downregulated antioxidants in the heart of the obese ZSF1 rats, which was augmented after PDE9A inhibition. From this data, it was concluded that PDE9A inhibition, potentially through augmenting the PRDX-5 antioxidant mechanism, improved coronary vascular rarefaction and vasodilator function, thereby restoring LV diastolic function in obese ZSF1 rats. T32HL155011 This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

The disintegrin and metalloproteinase 17 (ADAM17) exhibits α-secretase activity, whereby it can prevent the production of neurotoxic amyloid precursor protein-α (APP). ADAM17 is abundantly expressed in vascular endothelial cells and may act to regulate vascular homeostatic responses, including vascular growth and vasomotor function. We hypothesized that cerebrovascular ADAM17 plays a role in the pathogenesis of Alzheimer’s disease (AD). Here we found that 8-10 months old APP/PS1 mice (expressing transgenes for APP Swedish mutation and PSEN1 with L166P mutation) with significant cognitive deficits display a markedly reduced expression of ADAM17 in cerebral microvessels. Systemic delivery and AAV-mediated re-expression of ADAM17 in APP/PS1 mice improved cognitive functioning, without affecting β-amyloid plaque density. Moreover, we found that the endothelium-dependent vasodilator function of isolated and pressurized cerebral arteries in APP/PS1 mice was impaired, which was restored to normal levels by the ADAM17 re-expression. The cerebral microvascular network density in the APP/PS1 mice before or after ADAM17 re-expression was not affected. In addition, proteomic analysis identified several differentially expressed molecules involved in AD neurodegeneration and repair that were reversed by ADAM17 re-expression. Thus, we propose that a reduced ADAM17 expression in cerebral microvessels impairs vasodilator function, whereby it contributes to the development of cognitive dysfunction in APP/PS1 mice, and that ADAM17 can potentially be targeted for therapeutic intervention in AD. T32HL155011; 20PRE35211126 This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Accumulating epidemiological evidence shows that the incidence of multiple types of cancer, such as melanoma and GI cancers, is significantly higher in patients with heart failure with preserved ejection fraction (HFpEF). However, the mechanistic relationships between HFpEF and cancer development are poorly understood. Impaired angiogenesis, upregulation of endothelial adhesion molecules and chronic low-grade inflammation are common in HFpEF. Here we examined the role of endothelial cell-selective adhesion molecule (ESAM) in the progression of malignant melanoma in a HFpEF mouse model. Orthotopic melanoma xenografts (B16F10-Luc2) were used and implanted subcutaneously in mice underwent uninephrectomy and aldosterone infusion (UNX-Aldo with 1% high salt drinking water for 4 weeks) and tumor growth was monitored with bioluminescent imaging. We found that ESAM expression and serum levels were elevated in the UNX-Aldo mice, which was accompanied by enhanced melanoma growth and metastasis rate. ESAM promoted tube formation and increased phospho-Akt, phospho-PKCα and phospho-ERK1/2 in cultured endothelial cells. Flow cytometry analysis showed elevated number of tumor-associated neutrophils (CD11b+Ly6G+Ly6C-) and myeloid-derived suppressor cells (MDSCs, CD11b+F4/80-Gr-1+) in the UNX-Aldo mice. UNX-Aldo mice with genetic deletion of ESAM (ESAM KO) showed a reduced tumor growth and reduced number of tumor-associated neutrophils and MDSCs. Collectively, our results indicate that in HFpEF, a proinflammatory ESAM activation may act to promote tumor angiogenesis, neutrophil and MDSCs tumor infiltration, thereby mitigating a proper immune response and facilitate tumor growth. This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Effect of PLXDC2 knockdown on colony formation of radioresistant CAL27 and HN6 cells determined on Day 12 following 4 Gy IR exposure

Radiotherapy plays an essential role in the treatment of head and neck squamous cell carcinoma (HNSCC), yet radioresistance remains a major barrier to therapeutic efficacy. A better understanding of the predominant pathways determining radiotherapy response could help develop mechanism-informed therapies to improve cancer management. Here we report that radioresistant HNSCC cells exhibit increased tumor aggressiveness. Using unbiased proteome profiler antibody arrays, we identify that upregulation of c-Met phosphorylation is one of the critical mechanisms for radioresistance in HNSCC cells. We further uncover that radioresistance-associated HNSCC aggressiveness is effectively exacerbated by c-Met but is suppressed by its genetic knockdown and pharmacologic inactivation. Mechanistically, the resulting upregulation of c-Met promotes elevated expression of plexin domain containing 2 (PLXDC2) through activating ERK1/2-ELK1 signaling, which in turn modulates cancer cell plasticity by epithelial–mesenchymal transition (EMT) induction and enrichment of the cancer stem cell (CSC) subpopulation, leading to resistance of HNSCC cells to radiotherapy. Depletion of PLXDC2 overcomes c-Met–mediated radioresistance through reversing the EMT progress and blunting the self-renewal capacity of CSCs. Therapeutically, the addition of SU11274, a selective and potent c-Met inhibitor, to radiation induces tumor shrinkage and limits tumor metastasis to lymph nodes in an orthotopic mouse model. Collectively, these significant findings not only demonstrate a novel mechanism underpinning radioresistance-associated aggressiveness but also provide a possible therapeutic strategy to target radioresistance in patients with HNSCC.Significance:This work provides novel insights into c-Met-PLXDC2 signaling in radioresistance-associated aggressiveness and suggests a new mechanism-informed therapeutic strategy to overcome failure of radiotherapy in patients with HNSCC.

The primer sequences for qRT-PCR assays.

Physiological and pathological vascular remodeling is uniquely driven by mechanical forces from blood flow in which wall shear stress (WSS) mechanosensing by the vascular endothelium plays a pivotal role. This study aimed to determine the novel role for a disintegrin and metalloproteinase 17 (ADAM17) in impaired WSS mechanosensing, which was hypothesized to contribute to aging-associated abnormal vascular remodeling. Without changes in arterial blood pressure and blood flow rate, skeletal muscle resistance arteries of aged mice (30-month-old vs. 12-week-old) exhibited impaired WSS mechanosensing and displayed inward hypertrophic arterial remodeling. These vascular changes were recapitulated by in vivo confined, AAV9-mediated overexpression of ADAM17 in the resistance arteries of young mice. An aging-related increase in ADAM17 expression reduced the endothelial junction level of its cleavage substrate, junctional adhesion molecule-A/F11 receptor (JAM-A/F11R). In cultured endothelial cells subjected to steady WSS ADAM17 activation or JAM-A/F11R knockdown inhibited WSS mechanosensing. The ADAM17-activation induced, impaired WSS mechanosensing was normalized by overexpression of ADAM17 cleavage resistant, mutated JAM-A(V232Y) both in cultured endothelial cells and in resistance arteries of aged mice, in vivo. These data demonstrate a novel role for ADAM17 in JAM-A/F11R cleavage-mediated impaired endothelial WSS mechanosensing and subsequently developed abnormal arterial remodeling in aging. ADAM17 could prove to be a key regulator of WSS mechanosensing, whereby it can also play a role in pathological vascular remodeling in diseases. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s11357-021-00476-1.

Head and neck squamous cell carcinoma (HNSCC) remains a deadly disease despite concerted efforts to improve its diagnosis and treatment in recent decades. Metastasis of advanced HNSCC nearly always occurs first in neck lymph nodes before the development of distant metastasis. However, the development of preclinical animal models and therapeutic treatments for metastatic HNSCC is lagged from bench to clinic. In this protocol, we exemplify an orthotopic tongue tumor model that can recapitulate the cervical lymphatic metastases of HNSCC and the application to study the effect of novel saracatinib-loaded nanoparticles (Nano-Sar). By taking advantage of bioluminescence imaging (BLI), the present protocol reveals the strong anti-metastatic efficacy of Nano-Sar in the experimental setup. Collectively, the protocol with a novel metastatic mouse model shows great potential to evaluate treatments on metastatic diseases with the aid of bioluminescent technology.

The development of radioresistance in head and neck squamous cell carcinoma (HNSCC) remains a significant problem in cancer treatment, contributing to the lack of improvement in survival trends in the past decades. One of the clinical challenges is that radioresistance often promotes tumor aggressiveness. However, the underlying mechanisms and molecular determinants are largely unknown. We report here that radioresistance-associated HNSCC aggressiveness is effectively exacerbated by c-Met but can be suppressed by its genetic knockdown and pharmacological inactivation. Through unbiased RNAseq data, we further uncovered that the resulting upregulation of c-Met signaling increases the expression of PLXDC2. a critical gene associated with the tumor microenvironment. Mechanistically, PLXDC2 is upregulated in radioresistant HNSCC cells through c-Met mediated activation of ERK1/2-ELK1 signaling cascade. Most importantly, PLXDC2 modulates cancer cell plasticity by inducing epithelial-mesenchymal transition (EMT) and the emergence of cancer stem cell (CSC) subpopulation. Additionally, depletion of PLXDC2 overcomes c-Met-mediated radioresistance by reversing the EMT progress and blunting the self-renewal capacity of CSCs. Therapeutically, a combination of the c-Met selective inhibitor SU11274 with radiation remarkably induces tumor shrinkage and constrains tumor metastasis to lymph nodes in vivo. Our study is novel in being the first to explore the role and mechanism of the c-Met-PLXDC2 axis in radioresistance-associated HNSCC aggressiveness and the first to our knowledge to evaluate how to take advantage of blocking this signaling to overcome radioresistance in preclinical mouse models of HNSCC. Citation Format: Fanghui Chen, Liwei Lang, Chloe Shay, Georgia Chen, Nabil Saba, Yong Teng. Met confers radioresistance-associated aggressiveness through enhancing PLXDC2-mediated cancer stem cell plasticity [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 5793.

Numerous studies have clarified the effectiveness of paeoniflorin in the treatment of cholestasis. However, the therapeutic efficacy and mechanisms of action of paeoniflorin in intrahepatic cholestasis of pregnancy (ICP) were still unknown. This study aimed to investigate the molecular biological mechanisms of paeoniflorin against ICP by combining network pharmacology and metabolomics. The effects of paeoniflorin were investigated in the ICP rat model induced by 17α-ethinylestradiol, showing improvements in the liver indices, liver histopathological changes, bile flow rate, and serum levels of TBA and ALP. The underlying mechanisms and metabolic pathways of paeoniflorin were revealed by network pharmacology and untargeted metabolomics, showing that paeoniflorin exerted its curative effect against ICP-induced ferroptosis through PI3K/AKT and MAPK signalling pathways. In conclusion, paeoniflorin protected against ICP-induced liver injury through MAPK signaling pathways.

Background Two phase I studies assessed the pharmacokinetics of buprenorphine, its metabolite norbuprenorphine, and naloxone following administration of buprenorphine/naloxone sublingual tablets in Chinese participants. Methods In the first phase I, open-label, single ascending-dose (SAD) study, 82 opioid-naïve volunteers received a single buprenorphine/naloxone dose ranging from 2 mg/0.5 mg to 24 mg/6 mg while under naltrexone block. In a second phase I, open-label, multiple ascending-dose (MAD) study, 27 patients with opioid dependence in withdrawal received buprenorphine/naloxone doses of either 16 mg/4 mg or 24 mg/6 mg for 9 consecutive days. Serial blood samples were collected after a single dose (SAD study) and at steady-state (MAD study). Pharmacokinetic parameters were calculated using non-compartmental analysis. Safety assessments included adverse events monitoring and laboratory tests. Results The pharmacokinetic profiles of buprenorphine and naloxone were consistent between single- and multiple-dose studies. Peak plasma concentrations (Cmax) were reached early for buprenorphine (0.75–1.0 h) and naloxone (0.5 h), supporting rapid absorption. In the SAD study, increases in plasma exposures to buprenorphine and naloxone were less than dose proportional, in line with previous observations in Western populations. Buprenorphine-to-naloxone ratios for Cmax and area under the curve (AUC) were constant over the dose range investigated and also consistent with Western populations data. Steady state was reached within 7 days of daily dosing, with slight accumulation over repeated doses. No serious adverse events were observed. Conclusions The present data suggest that buprenorphine/naloxone pharmacokinetic profiles in Chinese participants are consistent, overall, with those in Western populations, supporting no differences in dosing. Clinical Trial Registration The protocols were registered on the official website of the China Food and Drug Administration (CFDA): http://www.chinadrugtrials.org.cn/; Registration numbers CTR20132963 (RB-CN-10-0012), CTR20140153 (RB-CN-10-0015). Electronic supplementary material The online version of this article (10.1007/s40268-019-0277-9) contains supplementary material, which is available to authorized users.

The identification and development of reagents that can limit preferential metabolic effectors in tumors as well as trigger unfavorable tumoral metabolic cues is of tremendous importance for suppressing malignancy. However, insufficient blocking of critical metabolic dependencies of cancer allows the development of metabolic bypasses, which significantly dampens therapeutic benefits of selective agents that target a single metabolic enzyme or pathway. We report here that head and neck squamous cell carcinoma (HNSCC) cells display strong addiction to glutamine. Devimistat, a novel lipoate analog, redirects cellular activity towards tumor-promoting glutaminolysis, leading to low anticancer efficacy in HNSCC cells. Mechanistically, devimistatinhibits the tricarboxylic acid cycle by blocking the enzyme activities of pyruvate dehydrogenase and alpha-ketoglutarate dehydrogenase, which upregulates GLS1 and eventually promotes the compensatory role of glutaminolysis in cancer cell survival. Most importantly, the addition of a GLS1 inhibitor CB-839 to devimistat treatment abrogates the metabolic dependency of HNSCC cells on glutamine, achieving a synergistic anticancer effect in glutamine-addicted HNSCC. These novel and significant findings could lay a scientific foundation for developing more effective treatments targeting the metabolic requirements of HNSCC. Citation Format: Liwei Lang, Fang Wang, Chloe Shay, Yonggang Ke, Nabil Saba, Yong Teng. Inhibition of glutaminolysis overcomes metabolic adaptation to devimistat treatment [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3026.

Targeting mitochondria as a cancer therapeutic strategy has attracted more and more attention in the recent years. ATAD3A is identified as a nuclear-encoded mitochondrial enzyme that regulates mitochondrial dynamics, cholesterol metabolism, and signal transduction. However, its impact and underlying regulatory mechanisms in cancers remain to be determined. We report here that ATAD3A is highly expressed in head and neck squamous cell carcinoma (HNSCC), where it plays a critical role in favoring tumor progression. Loss of ATAD3A expression dramatically incudes tumor regression in orthotopic tongue tumor-bearing mice, whereas gain of ATAD3A expression produces the opposite effects. Mechanistically, ATAD3A binds to ERK1/2 at HNSCC cell mitochondria in the presence of VDAC1, and this interaction is essential for the activation of mitochondrial ERK1/2 signaling. Most importantly, the ATAD3A-ERK1/2 signaling axis enhances HNSCC development in a Ras-independent manner and, thus, tumor suppression is more effectively achieved when ATAD3A and Ras signaling pathways are co-inhibited. These novel findings highlight the importance of activation of ATAD3A-ERK1/2 axis in cancer progression and suggest that combined targeting of ATAD3A and Ras signaling could potentiate anticancer activity for HNSCC treatment. Citation Format: Liwei Lang, Reid Loveless, Zhaohui Qin, Peng Qiu, Chloe Shay, Yong Teng. ATAD3A enhances head and neck cancer malignancy via mitochondrial ERK1/2 signaling [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 88.

One of the significant clinical challenges in head and neck cancer (HNC) treatment is a high rate of recurrence and metastasis after radiotherapy. Once tumor metastasis occurs following radiotherapy, the prognosis of HNC patients is extremely poor. However, the failure of radiotherapy for cancers is closely related to various factors, and the underlying mechanism remains unclear. We report here that c-Met is the most activated receptor tyrosine kinases in HNC cells once they develop radioresistance. Inactivation of c-Met dramatically attenuates the increased metastatic potential gained in HNC radioresistant cells, leading to tumor regression. Cancer stem cells (CSC) have been confirmed to be in a quiescent state in most established tumors. Mechanistically, the quiescent CSC population can be “awakened” by hyperactive c-Met signaling in radiation, which in turn initiates proliferation and differentiation to potentiate a series of malignant phenotypes. Most importantly, knockdown c-Met by shRNAs or inactivating it by c-Met inhibitor SU11274 reduces the population of “awakened” CSCs and suppresses metastasis, leading to increased sensitivity of HNC radioresistant cells to radiation. These novel findings highlight the importance of additional c-Met blockade in radiotherapy and suggest that combined targeting of c-Met and radiotherapy could be a promising strategy to improve the care for patients with HNC. Citation Format: Juan Dou, Liwei Lang, Georgia Chen, Nabil F. Saba, Yong Teng. Blockade of c-met signaling to circumvent radioresistance-associated metastasis in head and neck cancer [abstract]. In: Proceedings of the AACR Special Conference on the Evolutionary Dynamics in Carcinogenesis and Response to Therapy; 2022 Mar 14-17. Philadelphia (PA): AACR; Cancer Res 2022;82(10 Suppl):Abstract nr B038.

Coordination between the amplification of the fibroblast growth factor FGF19, overexpression of its corresponding receptor FGFR4, and hyperactivation of the downstream transmembrane enzyme β-klotho has been found to play pivotal roles in mediating tumor development and progression. Aberrant FGF19-FGFR4 signaling has been implicated in driving specific tumorigenic events including cancer cell proliferation, apoptosis resistance, and metastasis by activating a myriad of downstream signaling cascades. As an attractive target, several strategies implemented to disrupt the FGF19-FGFR4 axis have been developed in recent years, and FGF19-FGFR4 binding inhibitors are being intensely evaluated for their clinical use in treating FGF19-FGFR4 implicated cancers. Based on the established work, this review aims to detail how the FGF19-FGFR4 signaling pathway plays a vital role in cancer progression and why disrupting communication between FGF19 and FGFR4 serves as a promising therapeutic strategy for disrupting cancer progression.

The fibroblast growth factor 19 gene FGF19 has previously been reported to be amplified in several cancer types and encodes for a key autocrine signaler known to promote tumorigenic growth. Thus, it is imperative to understand which cancers are oncogenically addicted to FGF19 amplification as well as the role it serves in these cancer types. We report for the first time high FGF19 amplification in head and neck squamous cell carcinomas (HNSCC), which is associated with increased autocrine secretion of FGF19 and poor patient outcome in HNSCC. FGF19 amplification corresponded with constitutive activation of FGF receptor 4 (FGFR4)-dependent ERK/AKT-p70S6K-S6 signaling activation in HNSCC cells, and addition of human recombinant FGF19 could promote cell proliferation and soft agar colony formation in HNSCC cells with low FGF19 expression through activation of FGFR4 and downstream signaling cascades. In contrast, FGF19 knockout counteracts the observed effects in HNSCC cells carrying high endogenous FGF19, with knockout of FGF19 significantly suppressing tumor growth in an orthotopic mouse model of HNSCC. Collectively, this study demonstrates that FGF19 gene amplification corresponds with an increased dependency upon FGF19/FGFR4 autocrine signaling in HNSCC, revealing a therapeutic target for this cancer type.

Chronic periodontitis (CP) is a microbial dysbiotic disease linked to increased risk of oral squamous cell carcinomas (OSCCs). To address the underlying mechanisms, mouse and human cell infection models and human biopsy samples were employed. We show that the ‘keystone’ pathogen Porphyromonas gingivalis, disrupts immune surveillance by generating myeloid-derived dendritic suppressor cells (MDDSCs) from monocytes. MDDSCs inhibit CTLs and induce FOXP3 + Tregs through an anti-apoptotic pathway. This pathway, involving pAKT1, pFOXO1, FOXP3, IDO1 and BIM, is activated in humans with CP and in mice orally infected with Mfa1 expressing P. gingivalis strains. Mechanistically, activation of this pathway, demonstrating FOXP3 as a direct FOXO1-target gene, was demonstrated by ChIP-assay in human CP gingiva. Expression of oncogenic but not tumor suppressor markers is consistent with tumor cell proliferation demonstrated in OSCC-P. gingivalis cocultures. Importantly, FimA + P. gingivalis strain MFI invades OSCCs, inducing inflammatory/angiogenic/oncogenic proteins stimulating OSCCs proliferation through CXCR4. Inhibition of CXCR4 abolished Pg-MFI-induced OSCCs proliferation and reduced expression of oncogenic proteins SDF-1/CXCR4, plus pAKT1-pFOXO1. Conclusively, P. gingivalis, through Mfa1 and FimA fimbriae, promotes immunosuppression and oncogenic cell proliferation, respectively, through a two-hit receptor-ligand process involving DC-SIGN+hi/CXCR4+hi, activating a pAKT+hipFOXO1+hiBIM−lowFOXP3+hi and IDO+hi- driven pathway, likely to impact the prognosis of oral cancers in patients with periodontitis.

Background There is no consensus about the effective dosages of melatonin in cancer management, thus, it is imperative to fully understand the dose-dependent responsiveness of cancer cells to melatonin and the underlying mechanisms. Methods Head and neck squamous cell carcinoma (HNSCC) cells with or without melatonin treatment were used as a research platform. Gene depletion was achieved by short hairpin RNA, small interfering RNA, and CRISPR/Cas9. Molecular changes and regulations were assessed by Western blotting, quantitative RT-PCR (qRT-PCR), immunohistochemistry, and chromatin Immunoprecipitation coupled with qPCR (ChIP-qPCR). The therapeutic efficacy of FGF19/FGFR4 inhibition in melatonin-mediated tumor growth and metastasis was evaluated in orthotopic tongue tumor mice. Results The effect of melatonin on controlling cell motility and metastasis varies in HNSCC cells, which is dose-dependent. Mechanistically, high-dose melatonin facilitates the upregulation of FGF19 expression through activating endoplasmic stress (ER)-associated protein kinase RNA-like endoplasmic reticulum kinase (PERK)-Eukaryotic initiation factor 2 alpha (eIF2α)-activating transcription factor 4 (ATF4) pathway, which in turn promotes FGFR4-Vimentin invasive signaling and attenuates the role of melatonin in repressing metastasis. Intriguingly, following long-term exposure to high-dose melatonin, epithelial HNSCC cells revert the process towards mesenchymal transition and turn more aggressive, which is enabled by FGF19/FGFR4 upregulation and alleviated by genetic depletion of the FGF19 and FGFR4 genes or the treatment of FGFR4 inhibitor H3B-6527. Conclusions Our study gains novel mechanistic insights into melatonin-mediated modulation of FGF19/FGFR4 signaling in HNSCC, demonstrating that activating this molecular node confines the role of melatonin in suppressing metastasis and even triggers the switch of its function from anti-metastasis to metastasis promotion. The blockade of FGF19/FGFR4 signaling would have great potential in improving the efficacy of melatonin supplements in cancer treatment.

Background: Development of radioresistance in oral squamous cell carcinoma (OSCC) remains a significant problem in cancer treatment, contributing to the lack of improvement in survival trends in recent decades. Effective strategies to overcome radioresistance are necessary to improve the therapeutic outcomes of radiotherapy in OSCC patients. Methods: Cells and xenograft tumors were irradiated using the Small Animal Radiation Research Platform. AKT inhibitor capivasertib (AZD5363) was encapsulated into cathepsin B-responsible nanoparticles (NPs) for tumor-specific delivery. Cell viability was measured by alamarBlue, cell growth was determined by colony formation and 3D culture, and apoptosis was assessed by flow cytometry with the staining of Fluorescein isothiocyanate (FITC) Annexin V and PI. An orthotopic tongue tumor model was used to evaluate the in vivo therapeutic effects. The molecular changes induced by the treatments were assessed by Western blotting and immunohistochemistry. Results: We show that upregulation of AKT signaling is the critical mechanism for radioresistance in OSCC cells, and AKT inactivation by a selective and potent AKT inhibitor capivasertib results in radiosensitivity. Moreover, relative to irradiation (IR) alone, IR combined with the delivery of capivasertib in association with tumor-seeking NPs greatly enhanced tumor cell repression in 3D cell cultures and OSCC tumor shrinkage in an orthotopic mouse model. Conclusions: These data indicate that capivasertib is a potent agent that sensitizes radioresistant OSCC cells to IR and is a promising strategy to overcome failure of radiotherapy in OSCC patients.

With the development of new materials and technologies, it is possible to access gene function and drug metabolism using a three-dimensional (3D) cell culture system, which is more suitable for mimicking the in vivo microenvironment of cultured tumor cells ex vivo. SeedEZ is a novel and versatile tool that allows culturing of different types of cells with user convenience and in a desired sequence. This system provides a bridge between traditional 2D culture and animal experiments. Here, we provide two examples demonstrating how to evaluate cancer cell growth by the SeedEZ 3D scaffold and cancer cell invasion by the SeedEZ 3D ring, in order to promote understanding of the necessity of this novel cell culture system.

The functional relationship between apoptosis and autophagy in anticancer drug treatment is extremely complex, and the molecular machinery is obscure. This study aims to investigate the efficacy of CYT997, a novel microtubule-disrupting agent, in head and neck squamous cell carcinomas (HNSCCs) and complete the autophagy-apoptosis puzzle involved in drug action. We report here that CYT997 exhibits anticancer activity by triggering oxidative stress-associated apoptosis in HNSCC cells. Interestingly, upregulation of autophagy by mTOR-dependent pathways appears to have a cytoprotective role in preventing apoptosis by inhibiting CYT997-induced excessively high levels of reactive oxygen species (ROS). Blockade of autophagy by ATG7 depletion or addition of autophagy inhibitor hydroxychloroquine (HCQ) sensitizes HNSCC cells to CYT997 as evidenced by enhanced ROS-associated apoptosis. Moreover, HCQ exhibits a good synergism with CYT997 on induction of apoptosis in HNSCC xenografts without cytotoxicity, suggesting combined treatment of CYT997 with autophagy inhibitors would increase the anticancer efficacy of CYT997. These findings unveil the importance of ROS in crosstalk between autophagy and apoptosis in CYT997 treatment, raising concerns that genetic or pharmacologic blockade of autophagy should be considered in the design of new therapeutics for HNSCC. • CYT997 exhibits anticancer activity by induction of ROS-associated apoptosis. • mTOR-dependent cytoprotective autophagy prevents CYT997-induced apoptosis. • Blockade of autophagy augments CYT997 efficacy by enhanced ROS-associated apoptosis. • Combination of autophagy inhibitors with CYT997 is more effective against HNSCC.

The DNA binding protein and chromatin structural regulator DEK regulate many cellular processes. These include proliferation, differentiation, apoptosis, senescence, DNA repairing and the maintenance of stem cell phenotype. DEK is increasingly recognized as a crucial player in many steps of cancer initiation and progression, and is precisely regulated by abundant promoting and inhibiting factors directly or indirectly. DEK may serve as an architectural modulating protein to regulate the expression and function of multiple human genes in cancer cells. In this article we have reviewed the specificities and complexities of DEK in the regulation of transcription factors and global chromatin, including its biologic roles in malignant cells, and summarized the current research. The possible use of DEK as a diagnostic marker and drug target in the prevention or treatment of tumors is also discussed.

Head and neck squamous cell carcinoma (HNSCC) represents only 4% of all cancers in the US, but the adverse impact of this disease on health quality is dramatic. There has been little improvement in HNSCC treatment outcomes in decades, and a better understanding of the molecular biology of HNSCC is urgently needed to facilitate the design of anti-HNSCC therapies. The mitochondrial ATPase family AAA-domain containing protein 3A (ATAD3A) has been demonstrated as an oncogene in breast and lung cancer. However, nothing has been reported regarding its role in HNSCC. We report here that loss of ATAD3A expression inhibits HNSCC cell growth as evidenced by reduced cell proliferation and decreased the ability of colony formation and anchorage-independent growth in soft agar, whereas gain of ATAD3A expression counteracts the observed effects. Overexpression of a Walker A dead mutant of ATAD3A (ATAD3AK358) leads to a suppression in HNSCC cell growth and lipids synthesis, suggesting a potent dominant-negative effect of defective ATP-binding of ATAD3A. In line with the in vitro data, overexpression of ATAD3A facilities tumor growth in an orthotopic mouse model of HNSCC. In contrast, inhibition of ATAD3A function by knockout of ATAD3A or expression of ATAD3AK358 induces significant repression of tumor growth. These findings indicate that the mitochondrial protein ATAD3A plays an oncogenic role in HNSCC, and abrogating its ATP-binding ability may represent a promising regimen for better treatment of patients with HNSCC. Citation Format: Liwei Lang, Tiffany Lam, Alex Chen, Chole Shay, Yong Teng. The essential role of ATP binding to the mitochondrial protein ATAD3A in driving oncogenesis of head and neck squamous cell carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2406.

Background Src, an oncoprotein that drives progression of head and neck squamous cell carcinoma (HNSCC), is commonly hyperactivated in this disease. Unfortunately, the clinical benefit of targeting Src is significantly dampened in HNSCC patients, because the cytotoxic effects of anti-Src therapy and tumor resistance to it are less predictable. Thus, understanding the mechanism of tumor resistance to Src inhibition and seeking a way to overcome it are warranted. Methods Dual drug-loaded nanoparticles (NPs) were developed to co-deliver Src inhibitor saracatinib (AZD0530) and AKT inhibitor capivasertib (AZD5363) into the same population of tumor cells. An orthotopic tongue tumor model was generated to evaluate the in vivo therapeutic effects. Cell growth was determined by CellTiter-Glo® Luminescent Cell Viability Kit, colony formation, and 3D culture, and tumor growth was determined by bioluminescence and tumor size. The molecular changes induced by the treatments were assessed by Western blotting and immunohistochemistry. Results Capivasertib inactivated the AKT-S6 signaling and re-sensitized saracatinib-resistant HNSCC cells to saracatinib. Combination of capivasertib with saracatinib suppressed HNSCC growth more efficiently than either drug alone. Cathepsin B-sensitive NPs for co-delivering saracatinib and capivasertib significantly improved the efficacy of tumor repression without increasing side effects, which were due to highly specific tumor-targeting drug delivery system and synergistic anticancer effects by co-inactivation of AKT and Src in HNSCC cells. Conclusions Addition of AKT blockade improves anti-HNSCC efficacy of anti-Src therapy, and co-delivery of capivasertib and saracatinib by tumor-targeting NPs has the potential to achieve better treatment outcomes than the free drug combination.

Phosphofructokinase-1 (PFK-1), a rate-determining enzyme of glycolysis, is an allosteric enzyme that regulates the oxidation of glucose in cellular respiration. Glycolysis phosphofructokinase platelet (PFKP) is the platelet isoform and works as an important mediator of cell metabolism. Considering that PFKP is a crucial player in many steps of cancer initiation and metastasis, we reviewed the specificities and complexities of PFKP and its biological roles in human diseases, especially malignant tumors. The possible use of PFKP as a diagnostic marker or a drug target in the prevention or treatment of cancer is also discussed.

Background Metastatic lung cancer is a life-threatening condition that develops when cancer in another area of the body metastasizes, or spreads, to the lung. Despite advances in our understanding of primary lung oncogenesis, the biological basis driving the progression from primary to metastatic lung cancer remains poorly characterized. Methods Genetic knockdown of the particular genes in cancer cells were achieved by lentiviral-mediated interference. Invasion potential was determined by Matrigel and three-dimensional invasion. The secretion of matrix metalloproteinase 2 (MMP2) and MMP9 were measured by ELISA. Protein levels were assessed by Western blotting and immunohistochemistry. Protein-protein interactions were determined by immunoprecipitation. An experimental mouse model was generated to investigate the gene regulation in tumor growth and metastasis. Results Nck-associated protein 1 (NAP1/NCKAP1) is highly expressed in primary non-small-cell lung cancer (NSCLC) when compared with adjacent normal lung tissues, and its expression levels are strongly associated with the histologic tumor grade, metastasis and poor survival rate of NSCLC patients. Overexpression of NAP1 in lowly invasive NSCLC cells enhances MMP9 secretion and invasion potential, whereas NAP1 silencing in highly invasive NSCLC cells produces opposing effects in comparison. Mechanistic studies further reveal that the binding of NAP1 to the cellular chaperone heat shock protein 90 (HSP90) is required for its protein stabilization, and NAP1 plays an essential role in HSP90-mediated invasion and metastasis by provoking MMP9 activation and the epithelial-to-mesenchymal transition in NSCLC cells. Conclusions Our insights demonstrate the importance and functional regulation of the HSP90-NAP1 protein complex in cancer metastatic signaling, which spur new avenues to target this interaction as a novel approach to block NSCLC metastasis. Electronic supplementary material The online version of this article (10.1186/s13046-019-1124-0) contains supplementary material, which is available to authorized users.

Background A promising arsenal of histone deacetylase (HDAC)-targeted treatment has emerged in the past decade, as the abnormal targeting or retention of HDACs to DNA regulatory regions often occurs in many cancers. Head and neck squamous cell carcinoma (HNSCC) is one of the most aggressive malignancies worldwide associated with poor overall survival in late-stage patients. HDAC inhibitors have great potential to treat this devastating disease; however, few has been studied regarding the beneficial role of HDAC inhibition in anti-HNSCC therapy and the underlying molecular mechanisms remain elusive. Methods Cell migration and invasion were examined by wound closure and Transwell assays. Protein levels and interactions were assessed by Western blotting and immunoprecipitation. HDAC activity was measured with the fluorometric HDAC Activity Assay. Phospho-receptor tyrosine kinase (RTK) profiling was determined by the Proteome Profiler Human Phospho-RTK Array. Results ADP-ribosylation factor 1 (Arf1), a small GTPase coordinating vesicle-mediated intracellular trafficking, can be inactivated by HDAC inhibitors through histone acetylation-independent degradation of epidermal growth factor receptor (EGFR) in HNSCC cells. Mechanistically, high levels of Arf1 activity are maintained by binding to phosphorylated EGFR which is localized on HNSCC cell plasma membrane. Decreased EGFR phosphorylation is associated with reduced EGFR protein levels in the presence of TSA, which inactivates Arf1 and eventually inhibits invasion in HNSCC cells. Conclusions Our insights explore the critical role of EGFR-Arf1 complex in driving HNSCC progression, and demonstrate the selective action of HDAC inhibitors on this specific axis for suppressing HNSCC invasion. This novel finding represents the first example of modulating the EGFR-Arf1 complex in HNSCC by small molecule agents. Electronic supplementary material The online version of this article (10.1186/s13046-019-1080-8) contains supplementary material, which is available to authorized users.

Additional file 3: Figure S3. The effect of indicated treatment on other Src-related proteins (including STAT3, FAK and EGFR) in HNSCC cells.

Fibroblast growth factor receptor 4 (FGFR4), a tyrosine kinase receptor for FGFs, is involved in diverse cellular processes, including the regulation of cell proliferation, differentiation, migration, metabolism, and bile acid biosynthesis. High activation of FGFR4 is strongly associated with the amplification of its specific ligand FGF19 in many types of solid tumors and hematologic malignancies, where it acts as an oncogene driving the cancer development and progression. Currently, the development and therapeutic evaluation of FGFR4-specific inhibitors, such as BLU9931 and H3B-6527, in animal models and cancer patients, are paving the way to suppress hyperactive FGFR4 signaling in cancer. This comprehensive review not only covers the recent discoveries in understanding FGFR4 regulation and function in cancer, but also reveals the therapeutic implications and applications regarding emerging anti-FGFR4 agents. Our aim is to pinpoint the potential of FGFR4 as a therapeutic target and identify new avenues for advancing future research in the field.

Phosphofructokinase-1 (PFK-1), a rate-determining enzyme of glycolysis, is an allosteric enzyme that regulates the oxidation of glucose in cellular respiration. Glycolysis phosphofructokinase platelet (PFKP) is the platelet isoform and works as an important mediator of cell metabolism. Considering that PFKP is a crucial player in many steps of cancer initiation and metastasis, we reviewed the specificities and complexities of PFKP and its biological roles in human diseases, especially malignant tumors. The possible use of PFKP as a diagnostic marker or a drug target in the prevention or treatment of cancer is also discussed.

The ATPase family AAA-domain containing protein 3A (ATAD3A), a nuclear-encoded mitochondrial enzyme, is involved in diverse cellular processes, including mitochondrial dynamics, cell death and cholesterol metabolism. Overexpression and/or mutation of the ATAD3A gene have been observed in different types of cancer, associated with cancer development and progression. The dysregulated ATAD3A acts as a broker of a mitochondria-endoplasmic reticulum connection in cancer cells, and inhibition of this enzyme leads to tumor repression and enhanced sensitivity to chemotherapy and radiation. As such, ATAD3A is a promising drug target in cancer treatment.