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Abstract Background Small cell neuroendocrine carcinoma of the uterine cervix (SCNEC) is a rare cancer involving the human papilloma virus (HPV), and has few available treatments. The present work aimed to assess the feasibility of SOX2 and HPV statuses as predictive indicators of SCNEC prognosis. Methods The associations of SOX2 and/or high-risk (HR)-HPV RNA in situ hybridization (RISH) levels with clinicopathological characteristics and prognostic outcomes for 88 neuroendocrine carcinoma (NEC) cases were analyzed. Results Among these patients with SCNEC, SOX2, P16INK4A and HR-HPV RISH expression and SOX2/HR-HPV RISH co-expression were detected in 68(77.3%), 76(86.4%), 73(83.0%), and 48(54.5%), respectively. SOX2-positive and HR-HPV RISH-positive SCNEC cases were associated with poorer overall survival (OS, P = 0.0170, P = 0.0451) and disease-free survival (DFS, P = 0.0334, P = 0.0309) compared with those expressing low SOX2 and negative HR-HPV RISH. Alternatively, univariate analysis revealed that SOX2 and HR-HPV RISH expression, either separately or in combination, predicted the poor prognosis of SCNEC patients. Multivariate analysis revealed that the co-expression of SOX2 with HR-HPV RISH may be an independent factor of OS [hazard ratio = 3.597; 95% confidence interval (CI): 1.085–11.928; P = 0.036] and DFS [hazard ratio = 2.880; 95% CI: 1.199–6.919; P = 0.018] prediction in SCNEC. Conclusions Overall, the results of the present study suggest that the co-expression of SOX2 with HR-HPV RISH in SCNEC may represent a specific subgroup exhibiting remarkably poorer prognostic outcomes compared with the expression of any one marker alone.

Abstract Background To improve the efficiency of early diagnosis systems for cervical cancer, the use of cellular and viral markers for identifying precancerous lesions with a greater probability to progress to cancer has been proposed. Several cellular proteins and markers of oxidative DNA damage have been suggested as possible biomarkers of cervical carcinogenesis; however, they have not been evaluated together. In this study, we analyzed the expression of the cellular markers p16INK4a, Ki-67, CyclinE1, TOP2A/MCM2, and telomerase, as well as the DNA oxidative damage markers ROS and 8-OHdG. The analyses were performed in liquid-based cervical cytology samples or biopsies with premalignant lesions or cervical cancer diagnosis, with the purpose of selecting a panel of biomarkers that allow the identification of precursor lesions with greater risk of progression to cervical cancer. Methods We analyzed 1485 liquid-based cytology samples, including 239 non-squamous intraepithelial lesions (NSIL), 901 low-grade squamous intraepithelial lesions (LSIL), 54 high-grade squamous intraepithelial lesions (HSIL), and 291 cervical cancers (CC). The biomarkers were analyzed by immunocytochemistry and Human Papilloma Virus (HPV) genotyping with the INNO-LiPA genotyping Extra kit. Results We found that all tested cellular biomarkers were overexpressed in samples with high risk-HPV infection, and the expression levels increased with the severity of the lesion. TOP2A/MCM2 was the best biomarker for discriminating between LSIL and HSIL, followed by p16INK4a and cyclinE1. Statistical analysis showed that TOP2A/MCM2 provided the largest explanation of HSIL and CC cases (93.8%), followed by p16INK4a (91%), cyclin E1 (91%), Ki-67 (89.3%), and telomerase (88.9%). Conclusions We propose that the detection of TOP2A/MCM2, p16INK4a and cyclin E1 expression levels is useful as a panel of biomarkers that allow identification of cervical lesions with a higher risk for progression to CC with high sensitivity and precision; this can be done inexpensively, in a single and non-invasive liquid-based cytology sample.

Background: Small cell neuroendocrine carcinoma of the uterine cervix (SCNEC) is a rare cancer involving the human papilloma virus (HPV), and has few available treatments. The present work aimed to assess the feasibility of SOX2 and HPV statuses as predictive indicators of SCNEC prognosis. Methods: The associations of SOX2 and/or high-risk (HR)-HPV RNA in situ hybridization (RISH) levels with clinicopathological characteristics and prognostic outcomes for 88 neuroendocrine carcinoma (NEC) cases were analyzed. Results: Among these patients with SCNEC, SOX2, P16INK4A and HR-HPV RISH expression and SOX2/HR-HPV RISH co-expression were detected in 68(77.3%), 76(86.4%), 73(83.0%), and 48(54.5%), respectively. SOX2-positive and HR-HPV RISH-positive SCNEC cases were associated with poorer overall survival (OS, P = 0.0170, P = 0.0451) and disease-free survival (DFS, P = 0.0334, P = 0.0309) compared with those expressing low SOX2 and negative HR-HPV RISH. Alternatively, univariate analysis revealed that SOX2 and HR-HPV RISH expression, either separately or in combination, predicted the poor prognosis of SCNEC patients. Multivariate analysis revealed that the co-expression of SOX2 with HR-HPV RISH may be an independent factor of OS [hazard ratio = 3.597; 95% confidence interval (CI): 1.085–11.928; P = 0.036] and DFS [hazard ratio = 2.880; 95% CI: 1.199–6.919; P = 0.018] prediction in SCNEC. Conclusions: Overall, the results of the present study suggest that the co-expression of SOX2 with HR-HPV RISH in SCNEC may represent a specific subgroup exhibiting remarkably poorer prognostic outcomes compared with the expression of any one marker alone. [ABSTRACT FROM AUTHOR]

Abstract Background Cytotoxic chemotherapy can cure advanced germ cell tumors. Nevertheless, cancer treatment may induce cellular senescence and accelerate molecular aging. The aging process implies an increase of cells expressing p16 INK4a and changes in lymphocyte subpopulations. Our aim was to study the potential induction of premature immunosenescence in testicular cancer survivors (TCS) exposed to chemotherapy. Methods Case-control exploratory study of TCS treated with chemotherapy (≥3 BEP cycles, disease-free ≥3 months) compared with age matched healthy controls. Peripheral blood mononuclear cells were isolated, and lymphocyte subpopulations were analyzed by flow cytometry. CDKN2A/p16 INK4a expression in T cells was measured using qPCR. The percentage of lymphocyte subpopulations and the CDKN2A/p16 INK4a expression in TCS were compared with the control group using the Wilcoxon signed-rank test. Results We included 16 cases and 16 controls. The median age was 27 years (minimum 24, maximum 54) and the median time on surveillance was 26.5 months (minimum 3, maximum192). TCS had a lower percentage of total T cells and CD4+ T cells in total lymphocytes. Among the CD4+ T lymphocytes, TCS had less naïve CD4+ and increased memory CD4+ cells. Within the CD8+ T lymphocytes, TCS exhibited a decrease in the percentage of naïve cells and an increase in CD8 + CD45RA + CD57+ cells. TCS also exhibited decreased memory CD19+ B cells compared to the controls. The relative expression of CDKN2A/p16 INK4a in T cells was increased in TCS (mean 1.54; 95% CI of the mean: 1.074–2.005; p = 0.048). Conclusion In this exploratory study, TCS showed increased expression of CDKN2A/p16 INK4a and a lymphocyte phenotype that has been associated with immunosenescence. Further studies are warranted to define the clinical implications of these alterations in TCS.

Background: To improve the efficiency of early diagnosis systems for cervical cancer, the use of cellular and viral markers for identifying precancerous lesions with a greater probability to progress to cancer has been proposed. Several cellular proteins and markers of oxidative DNA damage have been suggested as possible biomarkers of cervical carcinogenesis; however, they have not been evaluated together. In this study, we analyzed the expression of the cellular markers p16INK4a, Ki-67, CyclinE1, TOP2A/MCM2, and telomerase, as well as the DNA oxidative damage markers ROS and 8-OHdG. The analyses were performed in liquid-based cervical cytology samples or biopsies with premalignant lesions or cervical cancer diagnosis, with the purpose of selecting a panel of biomarkers that allow the identification of precursor lesions with greater risk of progression to cervical cancer.Methods: We analyzed 1485 liquid-based cytology samples, including 239 non-squamous intraepithelial lesions (NSIL), 901 low-grade squamous intraepithelial lesions (LSIL), 54 high-grade squamous intraepithelial lesions (HSIL), and 291 cervical cancers (CC). The biomarkers were analyzed by immunocytochemistry and Human Papilloma Virus (HPV) genotyping with the INNO-LiPA genotyping Extra kit.Results: We found that all tested cellular biomarkers were overexpressed in samples with high risk-HPV infection, and the expression levels increased with the severity of the lesion. TOP2A/MCM2 was the best biomarker for discriminating between LSIL and HSIL, followed by p16INK4a and cyclinE1. Statistical analysis showed that TOP2A/MCM2 provided the largest explanation of HSIL and CC cases (93.8%), followed by p16INK4a (91%), cyclin E1 (91%), Ki-67 (89.3%), and telomerase (88.9%).Conclusions: We propose that the detection of TOP2A/MCM2, p16INK4a and cyclin E1 expression levels is useful as a panel of biomarkers that allow identification of cervical lesions with a higher risk for progression to CC with high sensitivity and precision; this can be done inexpensively, in a single and non-invasive liquid-based cytology sample. [ABSTRACT FROM AUTHOR]

Background: Chromosome 7 open reading frame 24 (C7orf24) was originally identified as a highly expressed protein in various types of cancer, and later shown to be a γ-glutamylcyclotransferase (GGCT). GGCT depletion in cancer cells has anti-proliferative effects in vitro and in vivo, and it is therefore considered a promising candidate as a therapeutic target. However, the cellular events induced by GGCT depletion remain unclear. Methods: GGCT was depleted by siRNA in MCF7, MDA-MB-231, PC3, A172, Hela, and LNCaP cells. Induction of cellular senescence was evaluated with senescence-associated β-galactosidase (SA-β-Gal) staining. Expression levels of p21WAF1/CIP1 and p16INK4A were assessed by qRT-PCR and Western blotting. Effects of simultaneous double knockdown of p21WAF1/CIP1 and p16INK4A together with GGCT on cell cycle regulation and cell growth was measured by flow cytometry, and trypan blue dye exclusion test. Results: We found that GGCT knockdown induces significant cellular senescence in various cancer cells. Cyclin dependent kinase inhibitor p21WAF1/CIP1 and/or p16INK4A were upregulated in all cell lines tested. Simultaneous knockdown of p21WAF1/CIP1 recovered the cell cycle arrest, attenuated cellular senescence induction, and rescued the subsequent growth inhibition in GGCT-silenced MCF7 breast cancer cells. In contrast, in GGCT silenced MDA-MB-231 breast cancer cells, GGCT depletion upregulated p16INK4A, which played a regulatory role in senescence induction, instead of p21WAF1/CIP1. Conclusions: Our findings demonstrate that induction of cellular senescence mediated by the upregulation of cyclin-dependent kinase inhibitors is a major event underlying the anti-proliferative effect of GGCT depletion in breast cancer cells, highlighting the potential of GGCT blockade as a therapeutic strategy to induce cellular senescence. [ABSTRACT FROM AUTHOR]

Background: Up to 50% of penile squamous cell carcinomas (pSCC) develop in the context of high-risk human papillomavirus (HR-HPV) infection. Most of these tumours have been reported to show basaloid differentiation and overexpression of tumour suppressor protein p16INK4a. Whether HPV-triggered carcinogenesis in pSCC has an impact on tumour aggressiveness, however, is still subject to research. Methods: In tissue specimens from 58 patients with surgically treated pSCC between 1995 and 2012, we performed p16INK4a immunohistochemistry and DNA extraction followed by HPV subtyping using a PCR-based approach. The results were correlated with histopathological and clinical parameters. Results: 90.4% of tumours were of conventional (keratinizing) subtype. HR-HPV DNA was detected in 29.3%, and a variety of p16INK4a staining patterns was observed in 58.6% of samples regardless of histologic subtype. Sensitivity of basaloid subtype to predict HR-HPV positivity was poor (11.8%). In contrast, sensitivity and specificity of p16INK4a staining to predict presence of HR-HPV DNA was 100% and 57%, respectively. By focussing on those samples with intense nuclear staining pattern for p16INK4a, specificity could be improved to 83%. Both expression of p16INK4a and presence of HR-HPV DNA, but not histologic grade, were inversely associated with pSCC tumour invasion (p = 0.01, p = 0.03, and p = 0.71). However, none of these correlated with nodal involvement or distant metastasis. In contrast to pathological tumour stage, the HR-HPV status, histologic grade, and p16INK4a positivity failed to predict cancer-specific survival. Conclusions: Our results confirm intense nuclear positivity for p16INK4a, rather than histologic subtype, as a good predictor for presence of HR-HPV DNA in pSCC. HR-HPV / p16INK4a positivity, independent of histological tumour grade, indicates a less aggressive local behaviour; however, its value as an independent prognostic indicator remains to be determined. Since local invasion can be judged without p16INK4a/HPV-detection on microscopic evaluation, our study argues against routine testing in the setting of pSCC. [ABSTRACT FROM AUTHOR]

Background: The polycomb group (PcG) family BMI1, acting downstream of the hedgehog (Hh) pathway, plays an essential role in the self-renewal of haematopoietic, neural, and intestinal stem cells, and is dysregulated in many types of cancer. Our recent report has demonstrated that Hh signalling activation can predict very earlier relapse of oesophageal cancers. As data were not available on the clinical role of BMI1 expression in oesophageal cancers after chemoradiotherapy (CRT), we analysed whether it could be also used to predict disease progression and prognosis in oesophageal cancer patients undergoing trimodality therapy of preoperative CRT and oesophagectomy. Methods: Expressions of BMI1 and p16INK4A, a downstream target of PcG, were analysed in 78 patients with histologically confirmed oesophageal squamous cell carcinoma (ESCC) after preoperative CRT by immunohistochemical staining. The association of BMI1 and p16INK4A expression with clinicopathologic characteristics was analysed by χ2-test. Survival analysis was carried out by the log-rank test using Kaplan-Meier method.Results: Among 78 ESCC patients, 24 patients (30.8%) showed BMI1 positivity, mainly localised in the nuclei of tumour cells. Patients harbouring BMI1-positive tumour cells showed significantly poorer prognoses than those without such cells or residual tumours (mean disease-free survival (DFS) time 16.8 vs 71.2 months; 3-yr DFS 13.3% vs 49.9%, P=0.002; mean OS time 21.8 vs 76.6 months; 3-yr OS 16.2% vs 54.9%, P=0.0005). There was no significant correlation between p16INK4A expression and BMI1 expression. Conclusions: Our study shows that BMI1 expression is a predictor of early relapse and poor prognosis in ESCC after CRT. These findings suggest that BMI1 signal activation might be involved in promoting cancer regrowth and progression after CRT, and might be indicative of emergence of ‘more aggressive’ cancer progenitor cells. [ABSTRACT FROM AUTHOR]

Background: Cytotoxic chemotherapy can cure advanced germ cell tumors. Nevertheless, cancer treatment may induce cellular senescence and accelerate molecular aging. The aging process implies an increase of cells expressing p16INK4a and changes in lymphocyte subpopulations. Our aim was to study the potential induction of premature immunosenescence in testicular cancer survivors (TCS) exposed to chemotherapy.Methods: Case-control exploratory study of TCS treated with chemotherapy (≥3 BEP cycles, disease-free ≥3 months) compared with age matched healthy controls. Peripheral blood mononuclear cells were isolated, and lymphocyte subpopulations were analyzed by flow cytometry. CDKN2A/p16INK4a expression in T cells was measured using qPCR. The percentage of lymphocyte subpopulations and the CDKN2A/p16INK4a expression in TCS were compared with the control group using the Wilcoxon signed-rank test.Results: We included 16 cases and 16 controls. The median age was 27 years (minimum 24, maximum 54) and the median time on surveillance was 26.5 months (minimum 3, maximum192). TCS had a lower percentage of total T cells and CD4+ T cells in total lymphocytes. Among the CD4+ T lymphocytes, TCS had less naïve CD4+ and increased memory CD4+ cells. Within the CD8+ T lymphocytes, TCS exhibited a decrease in the percentage of naïve cells and an increase in CD8 + CD45RA + CD57+ cells. TCS also exhibited decreased memory CD19+ B cells compared to the controls. The relative expression of CDKN2A/p16INK4a in T cells was increased in TCS (mean 1.54; 95% CI of the mean: 1.074-2.005; p = 0.048).Conclusion: In this exploratory study, TCS showed increased expression of CDKN2A/p16INK4a and a lymphocyte phenotype that has been associated with immunosenescence. Further studies are warranted to define the clinical implications of these alterations in TCS. [ABSTRACT FROM AUTHOR]

Background: Chromosome 7 open reading frame 24 (C7orf24) was originally identified as a highly expressed protein in various types of cancer, and later shown to be a γ-glutamylcyclotransferase (GGCT). GGCT depletion in cancer cells has anti-proliferative effects in vitro and in vivo, and it is therefore considered a promising candidate as a therapeutic target. However, the cellular events induced by GGCT depletion remain unclear., Methods: GGCT was depleted by siRNA in MCF7, MDA-MB-231, PC3, A172, Hela, and LNCaP cells. Induction of cellular senescence was evaluated with senescence-associated β-galactosidase (SA-β-Gal) staining. Expression levels of p21 WAF1/CIP1 and p16 INK4A were assessed by qRT-PCR and Western blotting. Effects of simultaneous double knockdown of p21 WAF1/CIP1 and p16 INK4A together with GGCT on cell cycle regulation and cell growth was measured by flow cytometry, and trypan blue dye exclusion test., Results: We found that GGCT knockdown induces significant cellular senescence in various cancer cells. Cyclin dependent kinase inhibitor p21 WAF1/CIP1 and/or p16 INK4A were upregulated in all cell lines tested. Simultaneous knockdown of p21 WAF1/CIP1 recovered the cell cycle arrest, attenuated cellular senescence induction, and rescued the subsequent growth inhibition in GGCT-silenced MCF7 breast cancer cells. In contrast, in GGCT silenced MDA-MB-231 breast cancer cells, GGCT depletion upregulated p16 INK4A , which played a regulatory role in senescence induction, instead of p21 WAF1/CIP1 ., Conclusions: Our findings demonstrate that induction of cellular senescence mediated by the upregulation of cyclin-dependent kinase inhibitors is a major event underlying the anti-proliferative effect of GGCT depletion in breast cancer cells, highlighting the potential of GGCT blockade as a therapeutic strategy to induce cellular senescence.