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The p14ARF-Hdm2-p53 signaling pathway targets tumor growth suppression, inhibits clonal proliferation, and maintains genome stability. It is inactivated in most human malignancies. The p14ARF protein is one of the main participants in the cascade. The aim of the study was to determine the prognostic value of the number of p14ARF-expressing tumor cells in biopsies of patients with diffuse large B cell lymphoma (DLBCL). Material and methods. The formalin-fixed paraffin embedded samples of tumor tissue of 104 patients with newly diagnosed DLBCL were included to the study. The relative number of cells expressing p14ARF was determined by immunohistochemical and morphometric methods. Differences in the content of p14ARF-positive tumor cells between groups of patients divided according to clinical and laboratory parameters were determined using the Mann–Whitney U test. Prediction of an unfavorable response to therapy was carried out using binary logistic regression with calculation of the odds ratio and 95 % confidence interval; variables were selected using Wald backward elimination method. The risk of an event occurring was calculated using Cox regression analysis. Results. The content of p14ARF-positive cells is higher in tumor biopsies of patients with high and high-intermediate risk according to the international prognostic index than in patients with low and lowintermediate risk. A relationship between an increase in the number of p14ARF-positive tumor cells, the absence of a complete response to first-line R-CHOP therapy, and an increased risk of death in patients with DLBCL has been established. Conclusions. The studied marker p14ARF can be used as an additional morphological predictor of the unfavorable course of DLBCL.

Prostate cancer (PCa) is characterized by a unique dependence on optimal androgen receptor (AR) activity where physiological androgen concentrations induce proliferation but castrate and supraphysiological levels suppress growth. This feature has been exploited in bipolar androgen therapy (BAT) for castrate resistant malignancies. Here, we investigated the role of the tumor suppressor protein p14ARF in maintaining optimal AR activity and the function of the AR itself in regulating p14ARF levels. We used a tumor tissue array of differing stages and grades to define the relationships between these components and identified a strong positive correlation between p14ARF and AR expression. Mechanistic studies utilizing CWR22 xenograft and cell culture models revealed that a decrease in AR reduced p14ARF expression and deregulated E2F factors, which are linked to p14ARF and AR regulation. Chromatin immunoprecipitation studies identified AR binding sites upstream of p14ARF. p14ARF depletion enhanced AR-dependent PSA and TMPRSS2 transcription, hence p14ARF constrains AR activity. However, p14ARF depletion ultimately results in apoptosis. In PCa cells, AR co-ops p14ARF as part of a feedback mechanism to ensure optimal AR activity for maximal prostate cancer cell survival and proliferation.

To investigate the protective effect of p14ARF in a nitric acid (NA) aerosol inhalation-induced bronchiolitis obliterans (BO) mouse model and its potential regulatory mechanism. A BO mouse model was established by NA aerosol inhalation. The expressions of p14ARF, phosphatidylinositol-3-kinase (PI3K), and protein kinase B (AKT) were detected by quantitative reverse transcription PCR (qRT-PCR) and western blot (WB). Hematoxylin (HE) staining, Masson staining, and periodic acid-Schiff (PAS) staining observed pulmonary histological changes. TdT-mediated dUTP nick end labeling (TUNEL) staining detected pulmonary cell apoptosis, and enzyme-linked immunosorbent assay (ELISA) measured matrix metalloproteinase-2 (MMP-2), MMP-9, tissue inhibitor of metalloproteinase-1 (TIMP-1), interleukon-6 (IL-6), and transforminh growth factor-β (TGF-β) levels in lung tissue and bronchoalveolar lavage fluid (BALF). The expressions of p14ARF, PI3K, and AKT showed a time gradient change, with a decrease trend (*P < 0.05 and **P < 0.01). Severe inflammatory infiltration and tracheal fibrosis were found in lung tissue in the modeling group (BO group) compared with the control group (Con group). The pH, PaO2, and PaO2/FiO2 values significantly reduced, while the PaCO2 value and the number of TUNEL-positive cells increased in BO group (P < 0.05). In addition, MMP-2, MMP-9, IL-6, and TGF-β levels remarkably increased, with an increase in the number of white blood cells, neutrophils, and lymphocytes in BO group (P < 0.05). Furthermore, p14ARF up-regulation reversed the trend of the aforementioned indexes in BO mice. p14ARF ameliorated the inflammatory response and airway remodeling in a BO mouse model via the PI3K/AKT pathway. [ABSTRACT FROM AUTHOR]

Cellular senescence is a major process affected by multiple signals and coordinated by a complex signal response network. Identification of novel regulators of cellular senescence and elucidation of their molecular mechanisms will aid in the discovery of new treatment strategies for aging-related diseases. In the present study, we identified human coilin-interacting nuclear ATPase protein (hCINAP) as a negative regulator of aging. Depletion of cCINAP significantly shortened the lifespan of Caenorhabditis elegans and accelerated primary cell aging. Moreover, mCINAP deletion markedly promoted organismal aging and stimulated senescence-associated secretory phenotype in the skeletal muscle and liver from mouse models of radiation-induced senescence. Mechanistically, hCINAP functions through regulating MDM2 status by distinct mechanisms. On the one hand, hCINAP decreases p53 stability by attenuating the interaction between p14ARF and MDM2; on the other hand, hCINAP promotes MDM2 transcription via inhibiting the deacetylation of H3K9ac in the MDM2 promoter by hindering the HDAC1/CoREST complex integrity. Collectively, our data demonstrate that hCINAP is a negative regulator of aging and provide insight into the molecular mechanisms underlying the aging process. [ABSTRACT FROM AUTHOR]

Purpose: The purpose of this study is to determine the relationship between APC gene mutations and subcellular differentiation levels of E2F1, P14ARF and KI67 proteins. Furthermore, if such connections can be used in the area of preventive health care. Materials and Methods: The 30 hours Immunohistochemistry protocol used, had samples preparation, antigen retrieval, background blocking, target detection and sample visualization. Samples were viewed and captured by light microscopy. Results: The conducted research concern 88 patients with a range age of 56 years. 57,7% had no tobacco addiction, 3,84% were obese and 19,23% had the tendency to consume alcohol. About 31% had at least one family member with a history of cancer. Intensity and positivity of the genes vary as seen in tables. Conclusions: By targeting specific and simultaneously multiple pathways based on molecular signatures, enables cases to be detected at an earlier stage, when there are greater chances of cure as treatment is more effective. A plan for early detection of cancer is a key component within an overall cancer control plan. An early diagnosis program is far more cheap and easy leading to appropriate treatments which finally reduce death rates and suffering due to cancer. [ABSTRACT FROM AUTHOR]

Current management of locally advanced rectal carcinoma (LARC) involves preoperative chemoradiotherapy (preCRT) before surgery. Despite improved local control rate, the response to preCRT of individual patients is variable and may reflect heterogeneous biological properties among tumors of the same clinical stage. Identifying novel molecular parameters with predictive and/or prognostic value is of great clinical importance for a personalized therapeutic approach. In this study, KRAS mutation status was analyzed by direct sequencing, while methylation-specific polymerase chain reaction (MSP) was used to examine p16INK4a and p14ARF gene methylation status in pretreatment tumor biopsies of 60 patients with LARC. The examined molecular changes of KRAS, p16INK4a and p14ARF genes were mutually independent (p16INK4a/KRAS, P=0.272; p14ARF/KRAS, P=0.923; p16INK4a/p14ARF, P=0.715). However, the simultaneous presence of p14ARF methylation and KRAS mutation was associated with a more frequent appearance of local recurrences and distant metastasis (P=0.027). Moreover, patients with the simultaneous presence of p16INK4a and p14ARF methylation and KRAS mutation had significantly shorter overall survival (P=0.011). The obtained results strongly suggest that combined analyses of examined genetic and epigenetic molecular alterations could contribute to the identification of LARC patient subgroups with more aggressive tumor behavior and worse disease outcome.

Background: It is well known that P16INK4A, P14ARF, P15INK4B mRNAs, and ANRIL lncRNA are transcribed from the CDKN2A/2B locus. LncRNA P14AS is a lncRNA transcribed from antisense strand of P14ARF promoter to intron-1. Our previous study showed that P14AS could upregulate the expression level of ANRIL and P16INK4A and promote the proliferation of cancer cells. Because polycomb group protein CBX7 could repress P16INK4A expression and bind ANRIL, we wonder whether the P14AS-upregulated ANRIL and P16INK4A expression is mediated with CBX7.Results: In this study, we found that the upregulation of P16INK4A, P14ARF, P15INK4B and ANRIL expression was induced by P14AS overexpression only in HEK293T and HCT116 cells with active endogenous CBX7 expression, but not in MGC803 and HepG2 cells with weak CBX7 expression. Further studies showed that the stable shRNA-knockdown of CBX7 expression abolished the P14AS-induced upregulation of these P14AS target genes in HEK293T and HCT116 cells whereas enforced CBX7 overexpression enabled P14AS to upregulate expression of these target genes in MGC803 and HepG2 cells. Moreover, a significant association between the expression levels of P14AS and its target genes were observed only in human colon cancer tissue samples with high level of CBX7 expression (n = 38, p < 0.05), but not in samples (n = 37) with low level of CBX7 expression, nor in paired surgical margin tissues. In addition, the results of RNA immunoprecipitation (RIP)- and chromatin immunoprecipitation (ChIP)-PCR analyses revealed that lncRNA P14AS could competitively bind to CBX7 protein which prevented the bindings of CBX7 to both lncRNA ANRIL and the promoters of P16INK4A, P14ARF and P15INK4B genes. The amounts of repressive histone modification H3K9m3 was also significantly decreased at the promoters of these genes by P14AS in CBX7 actively expressing cells.Conclusions: CBX7 expression is essential for P14AS to upregulate the expression of P16INK4A, P14ARF, P15INK4B and ANRIL genes in the CDKN2A/2Blocus. P14AS may upregulate these genes’ expression through competitively blocking CBX7-binding to ANRIL lncRNA and target gene promoters.

Abstract Although CDKN2A is well-known as a susceptibility gene for melanoma and pancreatic cancer, germline variants have also been anecdotally associated with a broader range of neoplasms including neural system tumors, head and neck squamous cell carcinomas, breast carcinomas, as well as sarcomas. The CDKN2A gene encodes for two distinct tumor suppressor proteins, p16INK4A and p14ARF, however, the independent association of germline alterations affecting these two proteins with cancer is under-appreciated. Here, we reviewed CDKN2A germline alterations reported among individuals and families with cancer in the literature, specifically addressing the cancer phenotypes in relation to the molecular consequence on p16INK4A and p14ARF. While melanoma is observed to associate with variants affecting both p16INK4A and p14ARF transcripts, it is noted that variants affecting p14ARF are more frequently observed with a heterogenous range of cancers. Finally, we reflected on the implications of this inferred genotype-phenotype association in clinical practice and proposed that clinical management of CDKN2A germline variant carriers should involve dedicated cancer genetics services, with multidisciplinary input from various healthcare professionals.

The goal of this study was to clarify the role of p16INK4a, p14ARF, and p53 protein expressions in carcinogenesis in squamous cell carcinomas of the lip. The expressions of the p53, p16INK4a, and p14ARF proteins were examined in 46 formaline-fixed paraffin embedded tissue specimens, which included 19 cases of squamous cell carcinoma of the lip, 14 cases of actinic cheilitis, and 13 cases of normal mucosa. Immunoreactivity in the peritumoral epithelium adjacent to squamous cell carcinomas was also evaluated. p16INK4a expression was increased in actinic cheilitis in comparison with normal mucosa (p=0.001). p14ARF expression progressively increased from normal mucosa to actinic cheilitis (p=0.001) and was observed to decrease significantly during the process of transition from actinic cheilitis to carcinoma (p=0.003). p53 values progressively increased from normal mucosa to actinic cheilitis (p=0.001) and carcinoma (p=0.008). A significant positive correlation was found between p14ARF and p53 in the peritumoral epithelium adjacent to carcinomas. Our findings indicated that p16INK4a and p14ARF immunohistochemistry does not determine whether or not actinic cheilitis has the potential to develop carcinoma. The p14ARF/p53 pathway is activated in the peritumoral epithelium adjacent to carcinoma; however, this activation would not be adequate to prevent carcinogenesis. [ABSTRACT FROM AUTHOR]

Background: Cervical cancer is the second most common cancer among women living in developing countries. Farnesoid X receptor (FXR) is a member of the nuclear receptor family, which regulates the development and proliferation of cancer. However, the role of and molecular mechanism by which FXR acts in cervical cancer are still unknown. Methods and results: The relationship between FXR and the proliferation of cervical cancer cell lines was detected by MTT and colony formation assays. Immunohistochemistry was used to detect the expression of FXR in cervical cancer tissue slides. Western blotting was used to detect the expression of p14ARF, mouse double minute 2 (MDM2) and p53 when FXR was overexpressed or siRNA was applied. Western blotting was used to detect the expression of MDM2 and p53 when pifithrin-α (PFT-α) was applied. FXR activation inhibited the proliferation of cervical cancer cell lines. FXR was significantly decreased in cervical squamous cell carcinoma, which was correlated with TNM stage, but not with metastasis. Overexpression of FXR activated the p14ARF-MDM2-p53 pathway. As a p53 inhibitor, PFT-α increased MDM2 in Lenti-vector cells, but had no effect on MDM2 in Lenti-FXR cells. Conclusions: FXR inhibits cervical cancer by upregulating the p14ARF-MDM2-p53 pathway. Activation of FXR may be a potential strategy for the treatment of cervical cancer. [ABSTRACT FROM AUTHOR]

Current management of locally advanced rectal carcinoma (LARC) involves preoperative chemoradiotherapy (preCRT) before surgery. Despite improved local control rate, the response to preCRT of individual patients is variable and may reflect heterogeneous biological properties among tumors of the same clinical stage. Identifying novel molecular parameters with predictive and/or prognostic value is of great clinical importance for a personalized therapeutic approach. In this study, KRAS mutation status was analyzed by direct sequencing, while methylation-specific polymerase chain reaction (MSP) was used to examine p16INK4a and p14ARF gene methylation status in pretreatment tumor biopsies of 60 patients with LARC. The examined molecular changes of KRAS, p16INK4a and p14ARF genes were mutually independent (p16INK4a/KRAS, P=0.272; p14ARF/KRAS, P=0.923; p16INK4a/p14ARF, P=0.715). However, the simultaneous presence of p14ARF methylation and KRAS mutation was associated with a more frequent appearance of local recurrences and distant metastasis (P=0.027). Moreover, patients with the simultaneous presence of p16INK4a and p14ARF methylation and KRAS mutation had significantly shorter overall survival (P=0.011). The obtained results strongly suggest that combined analyses of examined genetic and epigenetic molecular alterations could contribute to the identification of LARC patient subgroups with more aggressive tumor behavior and worse disease outcome. [ABSTRACT FROM AUTHOR]

p27Kip1 is known as a major cyclin‐dependent kinase inhibitor and a tumor suppressor, and often functionally hampered at protein level. p27 protein expression levels are frequently low in various cancers and negatively correlated with malignancy of cancer. However, in our previous study, we discovered that p27 overexpression does not inhibit the proliferation of two cancer cell lines due to a functional suppression of p27 by nucleophosmin isoform 1 (NPM1); that is, a qualitative, not quantitative, suppression of p27 function occurs in these cancer cell lines. To clarify the regulation of p27 in several types of cancer, we investigated p27 function in other cancer cell lines, based on proliferation assays in those cell lines carrying doxycycline‐inducible p27, and found that MDAH041 cells which express p14ARF, an antagonist of NPM1, show growth inhibition depending on p27 induction. Moreover, to investigate p27 function under anchorage‐independent culture conditions, we performed soft agar colony formation assay and observed that the colony formation of some cell lines carrying wild‐type p53, a major tumor suppressor, was inhibited depending on p27 induction. These results suggest that p27 function is regulated differentially among cancer cell types under anchorage‐dependent and anchorage‐independent culture conditions. [ABSTRACT FROM AUTHOR]

Anti-tumorigenic mechanisms mediated by the tumor suppressor p53, upon oncogenic stresses, are our bodies’ greatest weapons to battle against cancer onset and development. Consequently, factors that possess significant p53-regulating activities have been subjects of serious interest from the cancer research community. Among them, MDM2 and ARF are considered the most influential p53 regulators due to their abilities to inhibit and activate p53 functions, respectively. MDM2 inhibits p53 by promoting ubiquitination and proteasome-mediated degradation of p53, while ARF activates p53 by physically interacting with MDM2 to block its access to p53. This conventional understanding of p53-MDM2-ARF functional triangle have guided the direction of p53 research, as well as the development of p53-based therapeutic strategies for the last 30 years. Our increasing knowledge of this triangle during this time, especially through identification of p53-independent functions of MDM2 and ARF, have uncovered many under-appreciated molecular mechanisms connecting these three proteins. Through recognizing both antagonizing and synergizing relationships among them, our consideration for harnessing these relationships to develop effective cancer therapies needs an update accordingly. In this review, we will re-visit the conventional wisdom regarding p53-MDM2-ARF tumor-regulating mechanisms, highlight impactful studies contributing to the modern look of their relationships, and summarize ongoing efforts to target this pathway for effective cancer treatments. A refreshed appreciation of p53-MDM2-ARF network can bring innovative approaches to develop new generations of genetically-informed and clinically-effective cancer therapies.

Although CDKN2A is well-known as a susceptibility gene for melanoma and pancreatic cancer, germline variants have also been anecdotally associated with a broader range of neoplasms including neural system tumors, head and neck squamous cell carcinomas, breast carcinomas, as well as sarcomas. The CDKN2A gene encodes for two distinct tumor suppressor proteins, p16INK4A and p14ARF, however, the independent association of germline alterations affecting these two proteins with cancer is under-appreciated. Here, we reviewed CDKN2A germline alterations reported among individuals and families with cancer in the literature, specifically addressing the cancer phenotypes in relation to the molecular consequence on p16INK4A and p14ARF. While melanoma is observed to associate with variants affecting both p16INK4A and p14ARF transcripts, it is noted that variants affecting p14ARF are more frequently observed with a heterogenous range of cancers. Finally, we reflected on the implications of this inferred genotype-phenotype association in clinical practice and proposed that clinical management of CDKN2A germline variant carriers should involve dedicated cancer genetics services, with multidisciplinary input from various healthcare professionals. [ABSTRACT FROM AUTHOR]

Owing to its poor prognosis, the World Health Organization (WHO) lists lung cancer on top of the list when it comes to growing mortality rates and incidence. Usually, there are two types of lung cancer, small-cell lung cancer (SCLC) and non-small-cell lung cancer (NSCLC), which also includes adenocarcinoma, squamous cell carcinoma and large cell carcinomas. ARF, also known in humans as p14ARF and in the mouse as p19ARF, is a nucleolar protein and a member of INK4, a family of cyclin-independent kinase inhibitors (CKI). These genes are clustered on chromosome number 9p21 within the locus of CDKN2A. NSCLC has reported the role of p14ARF as a potential target. p14ARF has a basic mechanism to inhibit mouse double minute 2 protein that exhibits inhibitory action on p53, a phosphoprotein tumour suppressor, thus playing a role in various tumour-related activities such as growth inhibition, DNA damage, autophagy, apoptosis, cell cycle arrest and others. Extensive cancer research is ongoing and updated reports regarding the role of ARF in lung cancer are available. This article summarizes the available lung cancer ARF data, its molecular mechanisms and its associated signalling pathways. Attempts have been made to show how p14ARF functions in different types of lung cancer providing a thought to look upon ARF as a new target for treating the debilitating condition of lung cancer. [ABSTRACT FROM AUTHOR]

Bladder cancer (BlCa) is an extensively heterogeneous disease that leads to great variability in tumor evolution scenarios and lifelong patient surveillance, emphasizing the need for modern, minimally invasive precision medicine. Here, we explored the clinical significance of copy number alterations (CNAs) in BlCa. CNA profiling was performed in 15 patient-derived xenografts (PDXs) and validated in The Cancer Genome Atlas BlCa (TCGA-BLCA; n  = 408) and Lindgren et al. ( n  = 143) cohorts. CDKN2A copy number loss was identified as the most frequent CNA in bladder tumors, associated with reduced CDKN2A expression, tumors of a papillary phenotype, and prolonged PDX survival. The study's screening cohort consisted of 243 BlCa patients, and CDKN2A copy number was assessed in genomic DNA and cell-free DNA (cfDNA) from 217 tumors and 189 pre-treatment serum samples, respectively. CDKN2A copy number loss was correlated with superior disease-free and progression-free survival of non-muscle-invasive BlCa (NMIBC) patients. Moreover, a higher CDKN2A index ( CDKN2A/LEP ratio) in pre-treatment cfDNA was associated with advanced tumor stage and grade and short-term NMIBC progression to invasive disease, while multivariate models fitted for CDKN2A index in pre-treatment cfDNA offered superior risk stratification of T1/high-grade and EORTC high-risk patients, enhancing prediction of treatment outcome. CDKN2A copy number status could serve as a minimally invasive tool to improve risk stratification and support personalized prognosis in BlCa., Competing Interests: The authors declare no competing interests., (© 2024 The Author(s).)

p53 regulates the cellular response to genotoxic damage and prevents carcinogenic events. Theoretical and experimental studies state that the p53-Mdm2 network constitutes the core module of regulatory interactions activated by cellular stress induced by a variety of signaling pathways. In this paper, a strategy to control the p53-Mdm2 network regulated by p14ARF is developed, based on the pinning control technique, which consists into applying local feedback controllers to a small number of nodes (pinned ones) in the network. Pinned nodes are selected on the basis of their importance level in a topological hierarchy, their degree of connectivity within the network, and the biological role they perform. In this paper, two cases are considered. For the first case, the oscillatory pattern under gamma-radiation is recovered; afterward, as the second case, increased expression of p53 level is taken into account. For both cases, the control law is applied to p14ARF (pinned node based on a virtual leader methodology), and overexpressed Mdm2-mediated p53 degradation condition is considered as carcinogenic initial behavior. The approach in this paper uses a computational algorithm, which opens an alternative path to understand the cellular responses to stress, doing it possible to model and control the gene regulatory network dynamics in two different biological contexts. As the main result of the proposed control technique, the two mentioned desired behaviors are obtained.

Context: Urothelial carcinoma (UC) is one of the most frequent epithelial tumors worldwide. Aims: We aimed to investigate the protein expressions of caspase-8, p53, murine double minute 2 (mdm2), and p14ARF in nonmuscle UCs and to correlate the findings with clinicopathological characteristics. Settings and Design: A total of 50 patients who had pTa and pT1 tumors were analyzed. Subjects and Methods: The protein expressions of caspase-8, p53, mdm2, and p14ARF were analyzed by immunohistochemistry. Statistical Analysis Used: Chi-square test was done using SPSS version 16.0 (SPSS, Inc., Chicago, IL, USA). Results: Cytoplasmic caspase-8 expression was significantly higher in pT1 UCs while nuclear caspase-8 expression was significantly higher in pTa UCs (P = 0.005 and P = 0.011, respectively). Cytoplasmic caspase-8 expression was also higher in high-grade UCs (P = 0.035). The expression of p53, mdm2, and p14ARF was not also related with pathological stage or grade (P > 0.05 for all). The p14ARF expression was related with nuclear caspase-8 expression in most of the patients. Complete agreement among nonmuscle UCs for immunohistochemical expression of p14 and nuclear caspase-8 was seen in 41 cases, and the pairwise kappa agreement value was substantial (κ =0.614). The patients who had recurrence were positive for both p53 and mdm2 or either p53 or mdm2 (P = 0.025). Conclusions: These results suggested that the staining pattern of caspase-8 might be helpful for determining of the stages in nonmuscle UC. It was also showed that the expression status of p53 and mdm2 were related with the recurrence.

Background: Jab1 has been reported to regulate various proteins in signal transduction pathways and be implicated in carcinogenesis or tumor progression. However, the precise role and molecular mechanism of Jab1 in gastric tumorigenesis have not yet been fully elucidated. Methods: Jab1 staining in gastric cancer tissues and paired non-cancerous tissues was measured using tissue microarray (TMA) technology. The impact of Jab1 on tumor growth in vivo was analyzed using xenotransplantation experiments in Balb/c mice. The expression of Jab1 and p14ARF in gastric cancer cells was analyzed by western blot and confocal immunofluorescence. CCK-8 and cell cycle experiment were used to evaluate the cell proliferation. Ubiquitination assay was performed to validate whether ubiquitination is involved in Jab1-mediated p14ARF degradation. Results: The expression level of protein p14ARF was inversely correlated with the protein level of Jab1. Then, we investigated the mechanism that how Jab1 induced p14ARF depletion. Mechanistic studies showed that Jab1 induced ubiquitin-independent proteasomal p14ARF degradation in gastric cancer cells. Our data demonstrated that Jab1 protein was a vital upstream negative modulation factor of p14ARF, and Jab1 could promote cell proliferation and tumor growth via inhibiting the expression of p14ARF in vivo and in vitro. Moreover, silencing Jab1 protein expression declined tumor growth and further increased the apoptosis rate of gastric cancer cells. In further studies of gastric cancer specimens, we found the increased level of Jab1 protein shortened the overall survival. Conclusion: Jab1 is upstream of p14ARF and promote gastric cancer cell proliferation in vitro and in vivo. Furthermore, Jab1 decreased the expression of p14ARF though ubiquitination independent proteasomal degradation. Therefore, the connection of Jab1 and p14ARF may provide new methods for the treatment of gastric cancer. [ABSTRACT FROM AUTHOR]

p53 regulates the cellular response to genotoxic damage and prevents carcinogenic events. Theoretical and experimental studies state that the p53-Mdm2 network constitutes the core module of regulatory interactions activated by cellular stress induced by a variety of signaling pathways. In this paper, a strategy to control the p53-Mdm2 network regulated by p14ARF is developed, based on the pinning control technique, which consists into applying local feedback controllers to a small number of nodes (pinned ones) in the network. Pinned nodes are selected on the basis of their importance level in a topological hierarchy, their degree of connectivity within the network, and the biological role they perform. In this paper, two cases are considered. For the first case, the oscillatory pattern under gamma-radiation is recovered; afterward, as the second case, increased expression of p53 level is taken into account. For both cases, the control law is applied to p14ARF (pinned node based on a virtual leader methodology), and overexpressed Mdm2-mediated p53 degradation condition is considered as carcinogenic initial behavior. The approach in this paper uses a computational algorithm, which opens an alternative path to understand the cellular responses to stress, doing it possible to model and control the gene regulatory network dynamics in two different biological contexts. As the main result of the proposed control technique, the two mentioned desired behaviors are obtained. [ABSTRACT FROM AUTHOR]

The objective of the present study was to evaluate the epigenetic changes occurring in early stages of breast cancer. The present study investigated the methylation profile of the ATM, p14ARF and p16INK4a promoters in total blood and plasma cell-free DNA (cfDNA) from women with impalpable breast lesions compared with in total blood of a control cohort of women without breast lesions. The samples were evaluated using the methylation-specific PCR method. The Fisher's exact test was used to evaluate statistical significance between the methylation and clinical variables. A total of 111 women were evaluated, including 56 women with impalpable breast cancer (39/56 also had paired plasma cfDNA) and 55 women in the control cohort (55 blood DNA). For blood DNA from women with malignant impalpable breast lesions, p16INK4a exhibited the greatest percentage of methylation (48%), followed by ATM (37.5%) and p14ARF (27%) promoters, regardless of age variation. For plasma cfDNA, the methylation rates for ATM, p14ARF and p16INK4a were 26, 26 and 10%, respectively. The methylation rates for the blood DNA of controls were the lowest for ATM (9%), p14ARF (7%) and p16INK4a (7%). The women with impalpable breast lesions (benign and malignant lesions) exhibited the highest methylation rate, regardless of age, compared with the paired plasma cfDNA and controls. This epigenetic change was statistically significant for the promoters of ATM (P=0.009) and p16INK4a (P=0.001) (impalpable breast lesions vs. control). The present study demonstrated that epigenetic changes occurring in the ATM and CDKN2A genes detectable in liquid biopsy were associated with the development of impalpable breast lesions. [ABSTRACT FROM AUTHOR]

In this study, we identified P14 alternate reading frame (P14ARF) as a novel regulator of inflammation and vascularization in intervertebral disk degeneration (IVDD). We collected IVD tissues from IVDD patients and normal individuals for analysis of P14ARF expression. We also induced experimental IVDD by needle puncture injuries in the caudal intervertebral disks of Sprague- Dawley (SD) rats and achieved recombinant adenovirus-mediated P14ARF overexpression in experimental IVDD rats. Regulation relationships between P14ARF and tissue inhibitors of metalloproteinases- 3 (TIMP3) were confirmed in P14ARF-overexpressed and TIMP3-depleted nucleus pulposus (NP) cells. Tube formation in vitro was evaluated in coculture systems of human umbilical vein endothelial cells (HUVECs) and rat degenerated NP cells (DNPCs). Inflammatory response was assessed from levels of TNF-γ, IL-1, and IL-6 and neovascularization from expression of endothelial growth factor (VEGF). The P14ARF and TIMP3 were downregulated in degenerated IVD tissue derived from patients and experimental IVDD rats. Overexpressed P14ARF suppressed inflammatory cytokine levels and vascularization. There was decreased in vitro tube formation in response to P14ARF overexpression and TIMP3 elevation. Finally, attenuated inflammatory responses and suppression of VEGF were achieved by P14ARF-mediated promotion of TIMP3 in rat DNPCs. Taken together, the present study reveals that P14ARF/TIMP3 modulation of inflammatory response and vascularization in the context of IVDD highlights a potential target for future therapeutic strategies. [ABSTRACT FROM AUTHOR]

The treatment response of acute lymphoblastic leukemia (ALL) depends on the percentage of lymphoblasts, cytogenetic aberrations, and altered gene expression. The analysis of the gene expression is applicable for determination of risk stratification and prognosis of cancers. c-MYC, P14ARF, MDM2, and P53 play a vital role in cell survival through a functional network. This study aimed to investigate the expression of these genes, also their correlation with immunophenotypic subtypes of ALL and the percentage of blasts. Real-time PCR was performed for the expression analysis of P53, MDM2, c-MYC, and P14ARF in the bone marrow or peripheral blood samples of 52 ALL patients and 13 normal samples as controls. The morphological analysis and flow cytometry were carried out to examine the phenotypes and percentage of lymphoblasts. The decreased expression levels of P53 and MDM2 were seen in ALL patients compared with control group. In T cell subgroup of ALL the expression of P14ARF gene was more decreased among other subgroups. The expression of MDM2 was decreased in ALL patients who were under the age of 16. Based on our study, the interaction between P53 and MDM2 might be more complex and different from reports published in previous studies. Our findings showed that MDM2 is not negatively correlated with P53, at least in our samples. It can be very effective on the current and future studies to use different techniques for analysis of genome, transcriptome, and proteome in definitive risk stratification and prognosis determination. [ABSTRACT FROM AUTHOR]

Background/Aim: This research was aimed to evaluate the association between three selected single nucleotide polymorphisms (SNPs) within the CDKN2A (P14ARF) tumour suppressor gene and the incidence of endometrial cancer (EC) in postmenopausal women. Patients and Methods: The study included 194 postmenopausal women; 144 with EC and 50 non- cancer controls. Genotypes in P14ARF rs3088440, rs3731217 and rs3731245 polymorphisms were assayed using PCR-RFLP and confirmed by sequencing. Results: Regarding the rs3088440 polymorphism, CT, and CT-TT genotypes, were more prevalent among EC patients than in controls (OR=5.55, p=0.023, OR=5.29, p=0.027; and OR=2.92, p=0.023, respectively). The T allele within rs3088440 was more prevalent in EC females than in controls (χ²=4.7, p=0.030). Considering rs3731217, TG and TG-GG genotypes were less prevalent among EC (OR=0.34, p=0.024 or p=0.023; and OR=0.38, p=0.035, respectively). Conclusion: Polymorphisms in the CDKN2A gene are associated with EC in postmenopausal women. [ABSTRACT FROM AUTHOR]

Background: Endometrial cancer is a common gynecological malignancy with good prognosis in the early stages of the disease. The CpG island in the promoter region of tumor-suppressor genes are frequently methylated in various types of human cancers. In the present study, we have examined the methylation status of the p16INK4a and p14ARF genes in endometrial cancer and healthy endometrium with the aim to identify correlations between promoter hypermethylation, disease risk, and clinicopathological parameters. Methods: We collected 28 formalin fixed paraffin embedded samples and 26 blood samples from endometrial cancer patients and 22 controls. Methylation-specific PCR was applied to analyze the promoter methylation status of the p16INK4a and p14ARF genes in the studied population. The results were analyzed with SPSS software version 20. Results: There was a significant difference between the study groups and the presence of promoter CpG hypermethylation status in the p14 (P

Objective: This study aims to investigate the effects of CDKN2A (p16INK4A/p14ARF) over-expression on the proliferation and migration of human melanoma A375 cells. Methods: Melanoma tissues and pigmented nevi tissues were collected. Human melanoma A375 cells were transfected by CDKN2A (p16INK4A) and CDKN2A (p14ARF) over-expressing vectors and then assigned into blank, negative control (NC), p16INK4A and p14ARF groups. The expression of CDKN2A (p16INK4A) and CDKN2A (p14ARF) mRNA and protein was detected by qRT-PCR and Western blotting. CCK-8, flow cytometry and Transwell assays were applied to observe cell proliferation, the cell cycle and apoptosis, and migration and invasion, respectively. The model of subcutaneous xenografts in nude mice was established to measure cell growth in vivo. Results: Compared with pigmented nevi tissues, CDKN2A (p16INK4A) and CDKN2A (p14ARF) mRNA and protein expression were significantly decreased in melanoma tissues. CDKN2A (p16INK4A) and CDKN2A (p14ARF) over-expression inhibited proliferation, migration, invasion and progression from G0/G1 to S phase of A375 cells and xenograft tumor growth, but promoted apoptosis. Conclusion: Our study demonstrated that over-expression of CDKN2A (p16INK4A) and CDKN2A (p14ARF) suppressed proliferation and migration of human melanoma A375 cells.

Background/Aim: The prognostic role of USP10 in epithelial ovarian cancer has been studied in various human cancers. Our aim was to evaluate the clinical and pathological significance of USP10 in epithelial ovarian cancer. Materials and Methods: Immunohistochemical analyses of the expression of USP10 and p14ARF by using tissue microarrays were performed in 336 ovarian tumours and the data were compared with clinicopathological variables. We examined their level of DNA methylation around the putative transcriptional start site in 5' CpG islands in fresh frozen tissues and ovarian cancer cells. Results: Expression of USP10 and p14ARF was significantly lower in cancer tissues than in normal epithelium. Low USP10 expression and a combined USP10/p14ARF low expression were revealed to be independent prognostic factors. A high degree of methylation in USP10 and p14ARF CpG islands was found by methylation specific PCR analysis in cancer than in normal tissues and cells. Conclusion: Decreased expression of USP10 or combined USP10/p14ARF decreased expression is a strong indicator of poor prognosis in patients with ovarian cancer. [ABSTRACT FROM AUTHOR]

The article offers information regarding a study which aims at evaluating p14 expression patterns in nevi and melanoma, and this analysis identifies p14 as a candidate biomarker for melanoma diagnosis and prognosis. It mentions information on presence or absence of a particular protein within melanocytes. It presents information on clinical and histologic features of benign nevi, cutaneous melanoma, and metastatic melanoma.

The spindle assembly checkpoint (SAC) is a cellular surveillance mechanism that ensures the fidelity of chromosomes segregation. Reduced expression of some of its components weakens the SAC and induces chromosome instability and aneuploidy, which are both well-known hallmarks of cancer cells. Centromere protein-E (CENP-E) is a crucial component of the SAC and its function is to facilitate kinetochore microtubule attachment required to achieve and maintain chromosome alignment. The present study investigates the possible role of p14ARF as a controller of aneuploid cells proliferation. We used RNA interference to induce aneuploidy by partial depletion of CENP-E in human primary fibroblasts (IMR90) and in near diploid tumor cells (HCT116). In contrast to IMR90 aneuploid cell number, which was drastically reduced and leaned towards the WT condition, HCT116 aneuploid cell numbers were slightly decreased at later time points. This euploidy restoration was accompanied by increased p14ARF expression in IMR90 cells and followed ectopic p14ARF re-expression in p14ARF-null HCT116 cells. Collectively, our results suggest that hampering proliferation of aneuploid cells could be an additional role of the p14ARF tumor suppressor. [ABSTRACT FROM AUTHOR]

Genetic alterations in the INK4a/ARF (or CDKN2A) locus have been reported in many cancer types, including melanoma; head and neck squamous cell carcinomas; lung, breast, and pancreatic cancers. In melanoma, loss of function CDKN2A alterations have been identified in approximately 50% of primary melanomas, in over 75% of metastatic melanomas, and in the germline of 40% of families with a predisposition to cutaneous melanoma. The CDKN2A locus encodes two critical tumor suppressor proteins, the cyclin-dependent kinase inhibitor p16INK4a and the p53 regulator p14ARF. The majority of CDKN2A alterations in melanoma selectively target p16INK4a or affect the coding sequence of both p16INK4a and p14ARF. There is also a subset of less common somatic and germline INK4a/ARF alterations that affect p14ARF, while not altering the syntenic p16INK4a coding regions. In this review, we describe the frequency and types of somatic alterations affecting the CDKN2A locus in melanoma and germline CDKN2A alterations in familial melanoma, and their functional consequences in melanoma development. We discuss the clinical implications of CDKN2A inactivating alterations and their influence on treatment response and resistance.

Background/aim: Urothelial bladder cancer arises from the accumulation of multiple epigenetic and genetic changes. We aimed to investigate the specificity and sensitivity of gene-specific promoter methylation of CDH1 and p14ARF genes in the early diagnosis of bladder cancer and compare those with other diagnostic tests in our population. Patients and methods: In the current study, 65 patients with urothelial bladder cancer and 35 controls without any history of cancer were recruited. Methylation profiles of CDH1 and p14ARF genes from tumor and urine samples were determined by methylation-specific polymerase chain reaction method. Results: Methylation of CDH1 and p14ARF genes in tumor samples was 95.4% and 78.5%, respectively. The methylation frequencies were found to be 68.8% for CDH1 gene and 72.9% for p14ARF gene in urine samples. Sensitivities of CDH1, p14ARF and urine cytology were found to be 67.4%, 72.1% and 34.9%, respectively, while their specificities were 93.9%, 63.6% and 93.9%, respectively. Conclusion: Aberrant promoter methylation of CDH1 and p14ARF genes can be used to detect urothelial bladder cancer. In low-grade tumors, when compared with urine cytology, combined methylation analysis of CDH1 and p14ARF genes may not increase the sensitivity to identify malignant cells in urine samples. [ABSTRACT FROM AUTHOR]

ΔNp63α is finely and strictly regulated during embryogenesis and differentiation. ΔNp63α is the only p63 isoform degraded by the proteasome after Ubiquitin and SUMO (Small Ubiquitin-like MOdifier) conjugation. Here, we show that p63 ubiquitylation per se is not the signal triggering p63 proteasomal degradation. Taking advantage of natural ΔNp63α mutants isolated by patients with Split Hand and Foot Malformation IV syndrome, we found that SUMO and Ub modifications are not redundant and both are required to guarantee efficient ΔNp63α degradation. Here, we present evidence that sumoylation and ubiquitylation of ΔNp63α are strongly intertwined, and none of the two can efficiently occur if the other is impaired. [ABSTRACT FROM AUTHOR]

The human ubiquitin ligase HUWE1 has key roles in tumorigenesis, yet it is unkown how its activity is regulated. We present the crystal structure of a C-terminal part of HUWE1, including the catalytic domain, and reveal an asymmetric auto-inhibited dimer. We show that HUWE1 dimerizes in solution and self-associates in cells, and that both occurs through the crystallographic dimer interface. We demonstrate that HUWE1 is inhibited in cells and that it can be activated by disruption of the dimer interface. We identify a conserved segment in HUWE1 that counteracts dimer formation by associating with the dimerization region intramolecularly. Our studies reveal, intriguingly, that the tumor suppressor p14ARF binds to this segment and may thus shift the conformational equilibrium of HUWE1 toward the inactive state. We propose a model, in which the activity of HUWE1 underlies conformational control in response to physiological cues—a mechanism that may be exploited for cancer therapy.

Background: Endometrial cancer is a common gynecological malignancy with good prognosis in the early stages of the disease. The CpG island in the promoter region of tumor-suppressor genes are frequently methylated in various types of human cancers. In the present study, we have examined the methylation status of the p16INK4a and p14ARF genes in endometrial cancer and healthy endometrium with the aim to identify correlations between promoter hypermethylation, disease risk, and clinicopathological parameters. Methods: We collected 28 formalin fixed paraffin embedded samples and 26 blood samples from endometrial cancer patients and 22 controls. Methylation-specific PCR was applied to analyze the promoter methylation status of the p16INK4a and p14ARF genes in the studied population. The results were analyzed with SPSS software version 20. Results: There was a significant difference between the study groups and the presence of promoter CpG hypermethylation status in the p14 (P<0.0001) and p16 (P<0.05) genes. p14 hypermethylation in the blood samples was associated with depth of myometrial invasion in endometrial cancer (P=0.03). A significant association existed between p16 methylation in tissue with endometrial cancer grade (P=0.06). No statistically significant difference existed between thep16INK4a andp14ARF promoter hypermethylations in blood (P=0.177) and formalin fixed paraffin embedded (P=0.221) samples. An association existed between p16INK4a and p14ARF gene hypermethy-lations in blood and tissue with diabetes. Conclusion: Our results have confirmed that epigenetic mechanisms play an important role in endometrial cancer incidence. They can be utilized as prognostic biomarkers for endometrial cancer. The lack of a significant difference between the p16INK4a and p14ARF promoter hypermethylations in blood and formalin fixed paraffin embedded samples has indicated that methylation status of a blood sample can be an early, non-invasive diagnostic marker in endometrial cancer. [ABSTRACT FROM AUTHOR]

p14ARF, a tumor suppressor protein, encoded by the p16 tumor suppressor gene, has been reported to be associated with the clinicopathological features of lung cancer. However, the evaluated outcomes were inconsistent and remained inconclusive. In this study, we conducted a meta-analysis to clarify the significance of p14ARF expression in lung cancer pathogenesis. Materials and methods: Electronic databases, PubMed, Web of Knowledge, Embase, and CNKI, were retrieved to collect relevant articles with inclusion and exclusion criteria. Using Stata 12.0 software, 95% confidence intervals (CIs) and odds ratios (ORs) were calculated. Results: A total of 15 eligible case-control studies that evaluated the relationship between p14ARF expression and lung cancer were included in the meta-analysis. The results demonstrated that there were significant associations between p14ARF expression and the risk of non-smallcell lung cancer (NSCLC), lung adenocarcinoma, and lung squamous carcinoma (for NSCLC, OR =11.02, 95% CI =5.30-22.92; for lung adenocarcinoma, OR =7.28, 95% CI =3.92-13.50; and for lung squamous carcinoma, OR =14.40, 95% CI =2.83-73.24). In the stratified analysis based on race, significant associations between p14ARF expression and lung cancer risk were found in Chinese population and Caucasians (for Chinese population, OR = 7.02, 95% CI =4.48-11.00 and for Caucasians, OR =4.19, 95% CI =1.42-12.38). Furthermore, the expression of p14ARF was significantly associated with the TNM-stage of lung cancer in Chinese population (OR =2.07, 95% CI =1.38-3.10). Conclusion: p14ARF expression was significantly associated with the risk of lung cancer. In addition, the data of the meta-analysis showed that there was a significant correlation between p14ARF expression and the TNM-stage of lung cancer in Chinese population. [ABSTRACT FROM AUTHOR]