Aberrant Cytoplasmic p53 Expression and Its Correlation with TP53 Mutation Status and Functional Implications in Stage II and III Colorectal Cancer

Cytoplasmic p53 expression indicates a high frequency of TP53 abnormalities in gynaecological carcinoma. However, the implication of this expression in pulmonary neuroendocrine carcinoma (NEC) remains unclear. Thus, our study aimed to fill this research gap. Immunohistochemistry (IHC) of p53 was performed on 146 cases of resected small-cell lung carcinoma and large-cell NEC, and next-generation sequencing was conducted on cases showing cytoplasmic and wild-type p53 expression. IHC revealed overexpression in 57% of the cases (n = 83), complete absence in 31% (n = 45), cytoplasmic expression in 8% (n = 12) and wild-type expression in 4% (n = 6) of the cases. TP53 mutations were identified in nine of the 13 cases with available genetic analysis. The TP53 mutation rates in cases with cytoplasmic and wild-type p53 expression were 88% (seven of eight) and 40% (two of five), respectively. All seven cases showing cytoplasmic expression with TP53 mutations harboured loss-of-function type mutations: four had mutations in the DNA-binding domain, two in the nuclear localisation domain and one in the tetramerisation domain. Clinically, cases with cytoplasmic p53 expression had a poor prognosis similar to that in cases with p53 overexpression or complete absence. Cytoplasmic p53 expression in patients with pulmonary NEC suggests a high TP53 mutation rate, which is associated with a poor prognosis similar to that in patients with p53 overexpression or complete absence. This cytoplasmic expression should not be misidentified as a wild-type expression. This is the first report, to our knowledge, that demonstrates the implication of cytoplasmic p53 expression in pulmonary NEC.

Aberrant Cytoplasmic Expression of p16 Is a New Strong Prognostic Marker in Young MCL Patients Treated By Frontline Cytarabine-Based Immunochemotherapy, a Study from the Lysa Group, Nordic Lymphoma Group and the European MCL Network

Prognostic significance of cytoplasmic p53 oncoprotein in colorectal adenocarcinoma

p53 expression, DNA ploidy and S-phase fraction were analysed retrospectively in colorectal adenocarcinomas from 293 patients in whom the long-term outcome was known. The frequency of nuclear p53 staining was increased in non-diploid tumours (42%) when compared with diploid tumours (33%). Cytoplasmic p53 positive tumours were more common in the proximal colon (32%) than in the distal sites (21%). In univariate survival analysis, nuclear p53 and cytoplasmic staining were significantly associated with poor prognosis in patients with Dukes' A-C tumours. The patients showing both nuclear and cytoplasmic p53 staining had the poorest survival and the patients with tumours negative in both the nucleus and cytoplasm showed the best prognosis. The patients with tumours positive in the nucleus alone or in the cytoplasm alone presented an intermediate survival. In multivariate survival analyses, nuclear p53 expression, cytoplasmic p53 expression and DNA ploidy were prognostic indicators independent of Dukes' stage and each other. Further analysis suggested that the prognostic importance of cytoplasmic p53 expression was greater in diploid than in non-diploid tumours. We conclude that nuclear p53 expression, cytoplasmic p53 expression and DNA ploidy provide important prognostic information in colorectal adenocarcinomas.

Cytoplasmic localization of p27 (cyclin-dependent kinase inhibitor 1B/KIP1) in colorectal cancer: inverse correlations with nuclear p27 loss, microsatellite instability, and CpG island methylator phenotype

The present study was undertaken to evaluate the comparative expression of cell cycle regulators- p53, p21 and cdk2 in spontaneous canine tumours. Out of 46 cases diagnosed as tumour by histopathology, 19 were benign and 27 were malignant. Expression of p53, p21 and cdk2 was analyzed by employing immunohistochemical staining technique and their expression was detected in 84.62%, 69.23% and 69.23% cases, respectively. The percent expressions of p53, p21 and cdk2 showed variation among different histopathological types of tumours. Visceral/mucosal tumours exhibited higher expression of p53 when compared tocutaneous tumours. Expression of p21 was found to be 62.5% and 80% whereas cdk2 was 68.75% and 70% in skin and visceral/mucosal tumours, respectively. Analysis of individual cases for the expression and localization of these three biomarkers revealed seven different patterns. Out of 15 tumours which showed both p53 and p21 overexpression, 11 tumours showed cytoplasmic expression of p21 along with overexpression of cdk2, which indicated that cytoplasmic p21 was not able to perform its normal function of blocking the expression of cdk2. Overexpression of p53 along with cdk2 was observed in 5 cases where p21 expression was totally absent indicating overexpression of abnormal or mutated p53 which was not able to induce the expression of p21 leading to overexpression of cdk2. From present study it can be concluded that cytoplasmic or abnormal p21 expression is induced by either the abnormal/mutated p53 or any other pathway independent of p53.

Targeted Repression of TP53 Promotes Erythropoiesis in Del(5q) MDS and Overcomes Clinical Resistance to Lenalidomide

BackgroundPelvic floor disorders affect almost 50% of aging women. An important role in the pelvic floor support belongs to the levator ani muscle. The p27/kip1 (p27) protein, multifunctional cyclin-dependent kinase inhibitor, shows changing expression in differentiating skeletal muscle cells during development, and relatively high levels of p27 RNA were detected in the normal human skeletal muscles.MethodsBiopsy samples of levator ani muscle were obtained from 22 symptomatic patients with stress urinary incontinence, pelvic organ prolapse, and overlaps (age range 38–74), and nine asymptomatic women (age 31–49). Cryostat sections were investigated for p27 protein expression and type I (slow twitch) and type II (fast twitch) fibers.ResultsAll fibers exhibited strong plasma membrane (and nuclear) p27 protein expression. cytoplasmic p27 expression was virtually absent in asymptomatic women. In perimenopausal symptomatic patients (ages 38–55), muscle fibers showed hypertrophy and moderate cytoplasmic p27 staining accompanied by diminution of type II fibers. Older symptomatic patients (ages 57–74) showed cytoplasmic p27 overexpression accompanied by shrinking, cytoplasmic vacuolization and fragmentation of muscle cells. The plasma membrane and cytoplasmic p27 expression was not unique to the muscle cells. Under certain circumstances, it was also detected in other cell types (epithelium of ectocervix and luteal cells).ConclusionsThis is the first report on the unusual (plasma membrane and cytoplasmic) expression of p27 protein in normal and abnormal human striated muscle cells in vivo. Our data indicate that pelvic floor disorders are in perimenopausal patients associated with an appearance of moderate cytoplasmic p27 expression, accompanying hypertrophy and transition of type II into type I fibers. The patients in advanced postmenopause show shrinking and fragmentation of muscle fibers associated with strong cytoplasmic p27 expression.

Reduced nuclear p27 expression is associated with a poor outcome in various cancers, including non-small cell lung cancer (NSCLC). Cytoplasmic p27 expression was shown to be associated with an unfavorable response to chemotherapy and poor outcomes in some carcinomas, but it has not been well studied in NSCLC. Herein, p27 expression in 219 tumors surgically resected from NSCLC patients was evaluated by immunohistochemistry (IHC). The most common of p27 immunostaining in lung tumors was observed in the cytoplasm (N-/C+, 32%), followed by negative (N-/C-, 29%), nucleus (N+/C-, 24%), and nucleus plus cytoplasm (N+/C+, 15%). Kaplan-Meier and Cox regression models showed that p27 N-/C+ tumors exhibited the worst overall survival (OS) and relapse-free survival (RFS) among the four categories of tumors. Among 135 of 219 patients who received cisplatin-based chemotherapy, p27 N-/C+ tumors most commonly showed an unfavorable response to cisplatin-based chemotherapy, followed by p27 N-/C- tumors when p27 N+/C- tumors were used as a reference. IHC analysis for phosphorylated extracellular signal-regulated kinase (p-ERK) and Bcl-2 expression in the lung tumors was performed to test whether ERK activation could enhance p27 nuclear export and the expression of Bcl-2 to test whether ERK activation could enhance p27 nuclear export and Bcl-2 expression. The data showed that p-ERK expression was positively correlated with cytoplasmic p27 (N-/C+) and Bcl-2 expression in the lung tumors. Patients with high Bcl-2-expressing tumors treated with cisplatin-based chemotherapy showed unfavorable predictive values in a subset of this study population. Therefore, we suggest that cytoplasmic p27 (N-/C+) via ERK-activated Bcl-2 expression may predict an unfavorable response to cisplatin-based chemotherapy and poor outcomes in NSCLC.

Immunohistochemical Expression of MYH Protein Can Be Used to Identify Patients With MYH-Associated Polyposis

The cyclin-dependent kinase inhibitor p27 plays important roles in cell proliferation, cell motility, and apoptosis. Interestingly, the nuclear and cytoplasmic p27 exert opposite biological functions. In this study, we investigated the prognostic impact of subcellular p27 expression. We constructed melanoma tissue microarrays in a large series of melanoma patients, including 29 normal nevi, 52 dysplastic nevi, 270 primary melanomas, and 148 metastatic melanomas. The expression level of subcellular p27 in different stages of melanocytic lesions and its prognostic significance were evaluated. Compared with dysplastic nevi, nuclear p27 expression was remarkably reduced in primary melanomas and further reduced in metastatic melanoma (P < 0.001 for both), whereas cytoplasmic p27 expression is significantly increased from dysplastic nevi to primary melanomas (P = 0.032) and further increased in melanoma metastases (P = 0.037). Although loss of nuclear p27 expression is correlated with a worse 5-year survival of primary melanoma patients in Kaplan-Meier analysis (P = 0.046), it is not a prognostic factor by multivariate Cox regression analysis. On the contrary, Kaplan-Meier analysis showed that gain of cytoplasmic p27 was associated with a poor 5-year survival of metastatic melanoma patients (P < 0.001). Multivariate Cox regression analysis revealed that positive cytoplasmic p27 expression is an independent prognostic factor to predict metastatic melanoma patient outcome. Cytoplasmic p27 may serve as a promising prognostic marker for metastatic melanoma. Because there is no reliable prognostic marker for metastatic melanoma, our finding may have important clinical implications using cytoplasmic p27 as a prognostic biomarker for advanced melanoma.

&lt;div&gt;Abstract&lt;p&gt;&lt;b&gt;Background:&lt;/b&gt; The cyclin-dependent kinase inhibitor p27 plays important roles in cell proliferation, cell motility, and apoptosis. Interestingly, the nuclear and cytoplasmic p27 exert opposite biological functions. In this study, we investigated the prognostic impact of subcellular p27 expression.&lt;/p&gt;&lt;p&gt;&lt;b&gt;Methods:&lt;/b&gt; We constructed melanoma tissue microarrays in a large series of melanoma patients, including 29 normal nevi, 52 dysplastic nevi, 270 primary melanomas, and 148 metastatic melanomas. The expression level of subcellular p27 in different stages of melanocytic lesions and its prognostic significance were evaluated.&lt;/p&gt;&lt;p&gt;&lt;b&gt;Results:&lt;/b&gt; Compared with dysplastic nevi, nuclear p27 expression was remarkably reduced in primary melanomas and further reduced in metastatic melanoma (&lt;i&gt;P&lt;/i&gt; &lt; 0.001 for both), whereas cytoplasmic p27 expression is significantly increased from dysplastic nevi to primary melanomas (&lt;i&gt;P&lt;/i&gt; = 0.032) and further increased in melanoma metastases (&lt;i&gt;P&lt;/i&gt; = 0.037). Although loss of nuclear p27 expression is correlated with a worse 5-year survival of primary melanoma patients in Kaplan–Meier analysis (&lt;i&gt;P&lt;/i&gt; = 0.046), it is not a prognostic factor by multivariate Cox regression analysis. On the contrary, Kaplan–Meier analysis showed that gain of cytoplasmic p27 was associated with a poor 5-year survival of metastatic melanoma patients (&lt;i&gt;P&lt;/i&gt; &lt; 0.001). Multivariate Cox regression analysis revealed that positive cytoplasmic p27 expression is an independent prognostic factor to predict metastatic melanoma patient outcome.&lt;/p&gt;&lt;p&gt;&lt;b&gt;Conclusion:&lt;/b&gt; Cytoplasmic p27 may serve as a promising prognostic marker for metastatic melanoma.&lt;/p&gt;&lt;p&gt;&lt;b&gt;Impact:&lt;/b&gt; Because there is no reliable prognostic marker for metastatic melanoma, our finding may have important clinical implications using cytoplasmic p27 as a prognostic biomarker for advanced melanoma. &lt;i&gt;Cancer Epidemiol Biomarkers Prev; 20(10); 2212–21. ©2011 AACR&lt;/i&gt;.&lt;/p&gt;&lt;/div&gt;

&lt;div&gt;Abstract&lt;p&gt;&lt;b&gt;Background:&lt;/b&gt; The cyclin-dependent kinase inhibitor p27 plays important roles in cell proliferation, cell motility, and apoptosis. Interestingly, the nuclear and cytoplasmic p27 exert opposite biological functions. In this study, we investigated the prognostic impact of subcellular p27 expression.&lt;/p&gt;&lt;p&gt;&lt;b&gt;Methods:&lt;/b&gt; We constructed melanoma tissue microarrays in a large series of melanoma patients, including 29 normal nevi, 52 dysplastic nevi, 270 primary melanomas, and 148 metastatic melanomas. The expression level of subcellular p27 in different stages of melanocytic lesions and its prognostic significance were evaluated.&lt;/p&gt;&lt;p&gt;&lt;b&gt;Results:&lt;/b&gt; Compared with dysplastic nevi, nuclear p27 expression was remarkably reduced in primary melanomas and further reduced in metastatic melanoma (&lt;i&gt;P&lt;/i&gt; &lt; 0.001 for both), whereas cytoplasmic p27 expression is significantly increased from dysplastic nevi to primary melanomas (&lt;i&gt;P&lt;/i&gt; = 0.032) and further increased in melanoma metastases (&lt;i&gt;P&lt;/i&gt; = 0.037). Although loss of nuclear p27 expression is correlated with a worse 5-year survival of primary melanoma patients in Kaplan–Meier analysis (&lt;i&gt;P&lt;/i&gt; = 0.046), it is not a prognostic factor by multivariate Cox regression analysis. On the contrary, Kaplan–Meier analysis showed that gain of cytoplasmic p27 was associated with a poor 5-year survival of metastatic melanoma patients (&lt;i&gt;P&lt;/i&gt; &lt; 0.001). Multivariate Cox regression analysis revealed that positive cytoplasmic p27 expression is an independent prognostic factor to predict metastatic melanoma patient outcome.&lt;/p&gt;&lt;p&gt;&lt;b&gt;Conclusion:&lt;/b&gt; Cytoplasmic p27 may serve as a promising prognostic marker for metastatic melanoma.&lt;/p&gt;&lt;p&gt;&lt;b&gt;Impact:&lt;/b&gt; Because there is no reliable prognostic marker for metastatic melanoma, our finding may have important clinical implications using cytoplasmic p27 as a prognostic biomarker for advanced melanoma. &lt;i&gt;Cancer Epidemiol Biomarkers Prev; 20(10); 2212–21. ©2011 AACR&lt;/i&gt;.&lt;/p&gt;&lt;/div&gt;

Yes-associated protein (YAP) is a transcriptional coactivator regulated by autophagy that stimulates colorectal cancer (CRC) progression through activation of epithelial-mesenchymal transition (EMT), represented by tumor budding. The associations between these components in CRC are unknown. Archived surgically resected CRCs with known mismatch repair protein (MMR) status were retrieved (n=81; 2010 to 2016). Electronic medical records were reviewed for clinicopathologic variables including pathologic TNM stage and clinical stage. Tumor budding was graded according to consensus guidelines. Cytoplasmic and nuclear YAP and p62 (autophagy substrate) immunoreactivity were semiquantitatively scored within tumor samples. The Student t test, Fisher exact test, χ2 test, and Spearman correlation coefficient were performed with P<0.05 as a significance level. MMR proficiency (MMR-P) status correlated with high-grade tumor budding. The extent of cytoplasmic YAP staining and pathologic N stage was associated with tumor budding in multivariate analysis. Cytoplasmic YAP expression correlated with higher cytoplasmic p62 expression, suggesting an inverse correlation between autophagy activation and cytoplasmic YAP expression. Nuclear YAP expression correlated with pathologic N stage and clinical stage. A correlation between MMR-P status and tumor budding, combined with correlations between cytoplasmic YAP, tumor budding and p62 raise the possibility of 2 distinct neoplastic pathways concerning autophagy and YAP; one displaying relative activation of YAP and EMT, being commonly observed in MMR-P, and another with less active YAP and EMT, but active autophagy, being commonly seen in MMR-deficient CRC. Nuclear YAP staining could be useful in prognostication.

Herein we test the following hypotheses: (1) High-risk Human Papillomavirus (HR-HPV) may be involved in the etiology of mucoepidermoid carcinoma (MEC), and (2) The detection rate of HR-HPV in MEC has been increasing over time. Ninety-eight archival MEC specimens from three institutions spanning three decades were studied for HPV16/18 E6/E7 transcripts. RNA was extracted from formalin-fixed paraffin embedded specimens and HPV16/18 E6/E7 expression assessed by nested reverse transcription polymerase chain reaction (RT-PCR). A subset of MEC were also studied for MECT1-MAML2 fusion transcripts by nested RT-PCR and amplicon sequencing. The HPV expression data was validated by immunofluorescence (IF) with monoclonal HPV16/18 E6 antibody, PCR with the GP5+/6+ consensus primers, and sequencing of RT-PCR amplicons. HPV genome was localized by in-situ hybridization with the Ventana Inform HPVIII Family 16 probe. P16(INK4a) overexpression and aberrant p53 expression were assessed by immunohistochemistry. HPV16 E6/E7 transcripts were demonstrated in (29/98) 30% of MEC by RT-PCR. HPV18 E6/E7 transcripts were demonstrated in 13/98 (13%) of MEC by RT-PCR. Seven of 98 tumors (7%) demonstrated both HPV16/18. No significant association was found between HPV status and gender, age, and tumor site. All 13 HPV18+ MEC were diagnosed between 2001 and 2010, whereas 45 MEC diagnosed from 1977 to 2000 were negative for HPV18 (p = 0.002). By contrast, there was no significant difference with respect to HPV16 detection and date of diagnosis. All MEC that were positive for E6 protein were also HPV16/18 positive by RT-PCR. Sequencing a subset of RT-PCR amplicons confirmed HPV type- and region-specific sequences. PCR using GP5+/6+ consensus primers demonstrated HPV status concordance in 9 of 10 cases. DNA degradation was present in the last case; the RT-PCR amplicons were sequenced from this case which confirmed the presence of HPV type- and region-specific sequences. Strong (+4/+4) and diffuse (>50%) nuclear and cytoplasmic p16 expression was seen in 64% of MEC in the glandular regions, and 18% of MEC in the solid, squamoid regions. No correlation was seen between p16 expression and HPV status. Twenty-nine MEC (22 HPV+ and 7 HPV-negative) were selected for further evaluation for p53 expression. Strong aberrant nuclear p53 expression was present in only 2/22 HPV + MEC (9%, both Grade 3); no HPV-negative MEC demonstrated aberrant p53 expression. MECT1-MAML2 fusion transcripts were demonstrated in 23/37 (62%) MEC. No significant association was found between the presence of the MECT1-MAML2 fusion transcripts and tumor grade, HPV status, gender, era of diagnosis (2000 and earlier vs. 2001-2010) or tumor site. We demonstrate for the first time that transcriptionally active HPV16/18 is common to MEC. These findings were validated by demonstrating concordant results by separate PCR with consensus primers, and/or confirming the presence of HPV type- and region-specific sequences in the RT-PCR amplicons. We also visualized E6 viral oncoprotein and HPV genome within tumor cells. HR-HPV is thus potentially implicated in the pathogenesis of MEC. The frequency of HPV18 detection is significantly increased in MEC diagnosed after 2001, whereas we found no differences in the HPV16 detection rates per era of diagnosis.