P53-negative Tumors Research Articles

Colorectal cancer in patients seen at the teaching hospitals of Guadeloupe and Martinique: discrepancies, similarities in clinicopathological features, and p53 status

BackgroundIn Guadeloupe and Martinique, two French Overseas Departments, colorectal cancer (CRC) has become an essential public health issue. However, little is known about CRC characteristics and the p53 status in these populations, particularly in Guadeloupe, whereas certification of a cancer registry has been recently validated.MethodsThis was a descriptive retrospective study of 201 patients who, between 1995 and 2000, underwent surgery for CRC in the Guadeloupe Teaching Hospital (GlpeTH; 83 patients) and in the Martinique Teaching Hospital (MqueTH; 118 patients). The clinicopathological features and the p53 expression, evaluated with immunohistochemistry, were compared at the time of diagnosis. A relationship between these parameters and the p53 expression was also studied. Data were analysed, using the SPSS computer software version 17.0.ResultsNo statistical difference was found between the two groups of patients regarding age (p = 0.60), percentage of young patients (≤50 years; p = 0.94)), sex (p = 0.47), histological type (p = 0.073) and tumour sites (p = 0.65), although the GlpeTH patients were diagnosed with more distal colon cancers (54.2%) than the Mque TH patients (47.4%). By contrast, a significant difference was found regarding the tumour grade (p < 0.0001), the pTNM stage (p = 0.045) and the pT stage (p < 0.0001). Regarding p53 expression, solely for the MqueTH patients, nuclear expression was associated with pTNM, the percentage of p53 negative tumours increasing with the progression of the pTNM stages (p = 0.029).ConclusionsFor the first time, this study reveals discrepancies in clinicopathological features and in the p53 status between the two groups of patients. The GlpeTH patients were diagnosed with more moderated CRCs but with few CRCs at pTNM IV stage. By contrast, the MqueTH patients were diagnosed with more differentiated tumours, but with many more CRCs at pTNM IV stage. This paradox may be due to differences in tumour location (distal vs proximal), multiplicity of the genetic profiles of patients, or patients getting treatment elsewhere. Although our study is limited due to its small size, it emphasizes the originality of our results.

Epidemiology and survival of HPV‐related tonsillar carcinoma

The aim of this study was to determine the proportion of human papilloma virus (HPV)-positive cases in tonsillar carcinomas and investigate its development over the last decade. Further aim was to show the oncologic results in accord to HPV status and various treatment modalities. A retrospective study was conducted between 2000 and 2012 and included 275 patients treated for tonsillar carcinoma. P16 immunohistochemistry was used as a surrogate marker for HPV-associated carcinogenesis. A total of 101 (36.7%) patients proved to be p16 positive and 174 p16 negative. 80.2% of the p16-positive cases presented with T1-2 tumor. Of the early-stage patients, 79% of the p16-positive and 52.3% of the p16-negative presented with lymph node metastases. The percentage of p16-positive patients increased from 23.2% in the period 2005–2007 to 58.6% in the period 2010–2012 in the whole population and from 30.9% to 76.9% in T1-2 carcinomas. Early T-category p16-positive carcinomas had significantly better disease-specific survival (92.4% vs. 75.5%, P = 0.007) and overall survival (OS, 79.6% vs. 54.3%, P < 0.001) compared to p16-negative tumors. This study showed an increase in the percentage of p16-positive patients in tonsillar carcinoma from 23.2% in the years between 2005 and 2007 to 58.6% between 2010 and 2012. The majority (80.2%) of p16-positive patients presented with early T-category tumor but most of these (79.0%) had also lymph node metastases. Nevertheless, p16-positive patients had excellent oncologic results after surgery and adjuvant radiotherapy and could be considered for de-escalation of treatment.

Correlation of p16 status, hypoxic imaging using [18F]‐misonidazole positron emission tomography and outcome in patients with loco‐regionally advanced head and neck cancer

We investigated the relationship between hypoxia, human papillomavirus (HPV) status and outcome in head and neck squamous cell carcinoma. Patients with stage III and IV head and neck squamous cell carcinoma treated on phase I and II chemoradiation trials with 70-Gy radiation combined with tirapazamine/cisplatin or cisplatin/fluorouracil (5FU), hypoxic imaging using [18F]-misonidazole positron emission tomography and known HPV status (by p16 immunohistochemistry) were included in this sub-study. Separate analyses were conducted to consider the impact of tirapazamine on HPV-negative tumours in the phase II trial. Both p16-positive oropharyngeal tumours and p16-negative head and neck squamous cell carcinoma tumours had a high prevalence of tumour hypoxia; 14/19 (74%) and 35/44 (80%), respectively. The distribution of hypoxia (primary, nodal) was similar. On phase II, trial patients with p16-negative hypoxic tumours had worse loco-regional control with cisplatin and 5FU compared with tirapazamine and cisplatin (P < 0.001) and worse failure-free survival (hazard ratio = 5.18; 95% confidence interval, 1.98-13.55; P = 0.001). Only 1 out of 14 p16-positive patients on the phase II trial experienced loco-regional failure. Hypoxia, as assessed by [18F]-misonidazole positron emission tomography, is frequently present in both p16-positive and negative head and neck cancer. Further research is required to determine whether hypoxic imaging can be used to predict benefit from hypoxia-targeting therapies in patients with p16-negative tumours.

Abstract P6-05-27: p53, c-Myc and EGFR protein interactions confer worse prognosis in patients with triple negative breast cancer (TNBC)

Abstract Background - aim: TNBC is a heterogeneous disease comprising different molecular subtypes that are mostly defined by gene expression profiling and are related to patient outcome. Herein, we investigated the prognostic significance of molecular subtype-related, surrogate protein markers. Methods: Immunohistochemistry was applied on tissue microarrays containing a series of 330 centrally assessed TNBC from early high-risk breast cancer patients treated with anthracyclines and/or taxanes. The following markers were examined (in parentheses: cut-offs for low/high and negative/positive): Ki67 (14%); p53 (10%); c-Myc (10%); P-cadherin (10%); E-cadherin (10%); Claudin-7 (H-score median: 105); CK5 (1%); EGFR (1%); androgen receptor (AR [1%]). Results: The majority of tumors (269/327, 81.5%) were grade-III ductal carcinomas of no special type. Tumors were frequently positive for Ki67 (239/314, 76.1%), E-cadherin (193/271, 71.2%), P-cadherin (249/302, 82.5%) and expressed markers of the basal phenotype (CK5+ and/or EGFR+, 268/316, 84.8%). Grade-III, Ki67-high carcinomas and tumours positive for E- and P-cadherin were more prevalent among those of basal phenotype (Fisher's exact p&amp;lt;0.001). Tumors expressing low levels of Claudin-7 were infrequently positive for AR and EGFR proteins (p = 0.015 and p = 0.003, respectively). AR positivity was more frequent among CK5 negative, grade I-II, and Ki67-low tumors (p = 0.021, p = 0.05 and p = 0.011 respectively). When examined individually, none of these markers was related to patient outcome. An interaction was noticed between p53 and c-Myc immunopositivity (Wald's p = 0.028 for disease-free [DFS] and p = 0.005 for overall survival [OS]). Among patients with c-Myc negative tumors, patients with p53 positive tumors (n = 63) had shorter median DFS (46.8 months) and OS (56 months) (HR 1.9; 95%CI 1.0-3.9; p = 0.049 and HR 2.28; 95%CI 1.0-4.8; p = 0.031, respectively) as compared to those with p53 negative tumors (n = 56; median DFS 69 months, OS 70 months). Similarly, in the absence of p53, c-Myc expression correlated with shorter median OS (44.5 months) (HR 2.1; 95%CI 1.0-4.5; p = 0.038) but showed only a trend for shorter DFS as compared to c-Myc negative tumors. p53 protein status was a discriminating factor for EGFR negative tumors (Wald's p = 0.052), whereby p53 positive tumors (n = 54) were associated with reduced DFS as compared to p53 negative tumors (n = 68; median DFS: 47 vs. 68.8 months; HR 2.1; 95%CI 1.1-4.0; p = 0.025). Accordingly, EGFR immunopositivity conferred a trend for shorter DFS within p53 negative cases. Conclusions: In the current study, it was not possible to assign prognostic significance to individual proteins that have been introduced as surrogate markers for gene expression-based molecular subtypes in TNBC. By contrast, significant interactions involving the status of p53 and c-Myc or EGFR were observed. It seems that aberrant p53 adversely influences tumor behaviour in the absence of c-Myc and EGFR, while the opposite is also effective but to a lesser extent. These findings are hypothesis generating and need to be pursued in sufficiently powered studies for validating the apparently mutually exclusive adverse effects of p53, c-Myc and EGFR on tumor behaviour. Citation Information: Cancer Res 2013;73(24 Suppl): Abstract nr P6-05-27.

Ki-67 expression is increased in p16-expressing triple-negative breast carcinoma and correlates with p16 only in p53-negative tumors

Increased Ki-67 expression is associated with worse prognosis in patients with triple-negative breast carcinoma (TNBC); Ki-67 is widely used as a prognostic marker for TNBC patients. p16 and p53 are tumor suppressors. The status of p53 expression can divide TNBCs into 2 biologically distinct subgroups. The relationship of p16 expression with Ki-67 and its association with the status of p53 in TNBC patients have not been well characterized. In this study, we investigated p16 expression in 60 high-grade invasive TNBC cases and its relationship with Ki-67 in different groups of TNBCs and correlated p16 with Ki-67 in p53-positive and p53-negative subgroups. The tumors were immunolabeled for p16, Ki-67, and p53. Tissue microarrays were constructed with each tumor and adjacent normal breast tissue. Of the 60 tumors, 45 (75%) were found to have p16 expression. The triple-negative tumors had significantly higher p16 expression compared with paired normal ducts (P < .0001). Mean expression level of Ki-67 in p16-positive tumors was significantly higher than that in p16-negative tumors regardless of the status of p53 (P < .05). p16 expression positively correlated with Ki-67 (69.05% ± 7.23%) in the 22 p53-negative tumors (r = 0.739; P < .001). However, no correlation was found between p16 and Ki-67 (77.2% ± 3.83%) in the 38 p53-positive tumors (r = 0.157; P = .424). These findings suggest that p16 may play a role in the proliferation and aggressiveness of p53-negative TNBC and provide insights into the potential prognostic value of p16 as well as a better understanding of tumor biology related to the Rb/p16 pathway abnormalities.

Subcellular localization of Mdm2 expression and prognosis of breast cancer

Mouse double minute 2 (Mdm2) is a negative regulator of the tumor suppressor p53. The p53-Mdm2 pathway may play a role in cancer development and prognosis, although the role of p53-Mdm2 in breast cancer remains unclear. p53 and Mdm2 expressions were determined by immunohistochemistry of tissue microarrays of 865 breast cancer patients who underwent surgery. Clinicopathological characteristics and survival data were analyzed. Mdm2 expression was categorized into four groups: negative, cytoplasm positive, nucleus positive, and concurrent nuclear and cytoplasm positive (N+&C+). Negative, cytoplasm-positive, nucleus-positive, and N+&C+ expressions of Mdm2 were observed in 59.2, 10.9, 27.8, and 2.1 % of patients, respectively. The N+&C+ group was associated with larger tumor size, higher grade, negativity for estrogen and progesterone receptors, HER2 positivity, high Ki-67 index, p53 positivity, and triple negative breast cancer. p53-positive tumors showed poorer overall survival than p53-negative tumors. The nucleus-positive and N+&C+ groups showed poorer disease-free survival than the negative and cytoplasm-positive groups. In multivariate analysis, nuclear Mdm2 expression including the N+&C+ group was significantly related to poor prognosis. Concurrent nuclear and cytoplasmic Mdm2 expression was an independent prognostic factor in patients with breast cancer. Subcellular localization of Mdm2 expression should be considered in the evaluation of Mdm2 in breast cancer.

Immunohistochemical investigation of the angiogenic proteins VEGF, HIF-1α and CD34 in invasive ductal carcinoma of the breast

The expression of prognostic markers in cancer has become important in diagnostic routine and research. A high mitotic rate compromises the individual cell access to oxygen and nutrients, due to reduced blood supply. Cells undertake adaptive measures such as vascular endothelial growth factor (VEGF), expressed under the control of hypoxia-inducible factor-1α (HIF-1α). CD34 is an endothelial marker which can show the presence and distribution of blood vessels. This study evaluated the presence and relative expression of VEGF, HIF-1α and CD34 using immunohistochemistry of 60 breast tumors and double staining, correlating the findings with clinical and pathological variables. High VEGF expression was correlated with low cell proliferation, lymph node-negative, smaller tumor size and patients not receiving hormone therapy. High HIF-1α expression predominated in younger (<50-year) patients, subjected to neo-adjuvant therapy and in p53-negative tumors. Absence of metastasis, radiotherapy or hormone treatment, and estrogen receptor (ER)-positive tumors showed high CD34 immunoreactivity. We suggest that the angiogenic factors VEGF, HIF-1α and CD34 are important in breast cancer progression and their abundance in breast tumors has prognostic and predictive value.

OP198: HPV alters the angiogenic profile in head and neck cancers

Purpose Head and neck cancers (HNC) are aggressive tumours. Expression of Human Papilloma Virus (HPV)-derived protein p16 in HNC is known to correlate with a better prognosis than p16-negative tumours. Angiogenesis is an important factor in tumour progression. Our aim was to dissect the impact of HPV on angiogenic factors in HNC. Methods 18 newly diagnosed HNC patients and controls were analysed in this study. A panel of 11 pro- and anti-angiogenic factors was quantified in serum samples of HNC patients and controls using multiplex ELISA and correlated to the p16 expression in the tumour tissue. Tumour tissue was tested using immunohistochemistry for the expression of angiogenic factors and co-localisation performed to identify the predominant cellular source of angiogenic factors. Results All patients had higher circulating levels of the antiangiogenic factor endostatin compared to healthy donors. Interestingly, higher levels of the proangiogenic factors angiopoetin, placental growth factor (PlGF) and vascular endothelial growth factor (VEGF) were noted in patients who did not express p16 compared to those who did. Tumour associated macrophages appear to be a major cellular source of these angiogenic factors and work is underway to analyse the expression of angiogenic factors in HPV-positive and negative tumour tissue. Conclusions HNC patients that lack p16 expression have a different pattern of angiogenic factors with increased proangiogenic factors angiopoietin, PlGF and VEGF. This may confer a more aggressive phenotype to p16-negative tumours. Better understanding of the mechanisms by which HPV induces changes in tumour angiogenesis may guide the development of novel therapeutic targets.

Human Papillomavirus, p16, and Epidermal Growth Factor Receptor Biomarkers and CT Perfusion Values in Head and Neck Squamous Cell Carcinoma

Head and neck squamous cell carcinoma tumors positive for laboratory biomarkers hrHPV and p16 and negative for EGFR often respond better to nonsurgical organ-preservation therapy than hrHPV-negative, p16-negative, and EGFR overexpressing tumors. CTP has been shown to distinguish which locally advanced head and neck squamous cell carcinomas will respond to induction chemotherapy or chemoradiation. Our purpose was to determine whether a relationship exists between CTP measures and the expression of these laboratory biomarkers, because both appear to separate head and neck squamous cell carcinoma tumors into similar groups. We conducted an institutional review board-approved, Health Insurance Portability and Accountability Act-compliant retrospective review of head and neck CTP in 25 patients with locally advanced head and neck squamous cell carcinoma who had signed informed consent. Eight women and 17 men, 41-80 years of age, constituted a pretreatment group of 18 patients and a palliative group of 7 patients. Tumor biopsy samples were analyzed for overexpression of hrHPV, p16, and EGFR. The hrHPV, p16, and EGFR status of the tumors was correlated with CTP parameters (MTT, BV, BF, CP) by using the Wilcoxon evaluation and Fischer exact test. There were significantly lower CP values in pretreatment tumors overexpressing EGFR (P = .04). CP values ≤17.23 were significantly correlated with EGFR overexpression (P = .015). A trend toward higher CP values was present in hrHPV-positive and p16-overexpressing pretreatment tumors (P = .14). A significant correlation exists between CTP measures and EGFR overexpression in head and neck squamous cell carcinomas, suggesting an association between certain imaging findings and molecular biomarkers. These results may be related to a tumor cell survival mechanism linking perfusion and biomarker expression.

Influence of anthropometric factors on tumour biological characteristics of colorectal cancer in men and women: a cohort study

BackgroundObesity is a well established risk factor of colorectal cancer (CRC), but how body size influences risk of colorectal cancer defined by key molecular alterations remains unclear. In this study, we investigated the relationship between height, weight, body mass index (BMI), waist- and hip circumference, waist-hip ratio (WHR) and risk of CRC according to expression of beta-catenin, cyclin D1, p53 and microsatellite instability status of the tumours in men and women, respectively.MethodsImmunohistochemical expression of beta-catenin, cyclin D1, p53 and MSI-screening status was assessed in tissue microarrays with tumours from 584 cases of incident CRC in the Malmö Diet and Cancer Study. Six anthropometric factors: height, weight, BMI, waist- and hip circumference, and WHR were categorized by quartiles of baseline measurements and relative risks of CRC according to expression of beta-catenin, cyclin D1, p53 and MSI status were calculated using multivariate Cox regression models.ResultsHigh height was associated with risk of cyclin D1 positive, and p53 negative CRC in women but not with any investigative molecular subsets of CRC in men. High weight was associated with beta-catenin positive, cyclin D1 positive, p53 negative and microsatellite stable (MSS) tumours in women, and with beta-catenin negative and p53 positive tumours in men. Increased hip circumference was associated with beta-catenin positive, p53 negative and MSS tumours in women and with beta-catenin negative, cyclin D1 positive, p53 positive and MSS tumours in men. In women, waist circumference and WHR were not associated with any molecular subsets of CRC. In men, both high WHR and high waist circumference were associated with beta-catenin positive, cyclin D1 positive and p53 positive tumours. WHR was also associated with p53 negative CRC, and waist circumference with MSS tumours. High BMI was associated with increased risk of beta-catenin positive and MSS CRC in women, and with beta-catenin positive, cyclin D1 positive and p53 positive tumours in men.ConclusionsFindings from this large prospective cohort study indicate sex-related differences in the relationship between obesity and CRC risk according to key molecular characteristics, and provide further support of an influence of lifestyle factors on different molecular pathways of colorectal carcinogenesis.

P53 and Expression of Immunological Markers May Identify Early Stage Thyroid Tumors

Background. Besides its major role in cell proliferation, DNA repair, and apoptosis, functional p53 protein is involved in the induction of antitumor cytotoxic-T-cell activity against carcinoma cells. We aimed to investigate p53 and immune cell markers utility as diagnostic and prognostic markers of differentiated thyroid cancer (DTC). Methods. ACIS-III system was used to evaluate p53 and immune cell markers including tumor-associated macrophages (TAM); CD68 and tumor-infiltrating lymphocytes (TIL) subsets such as CD3, CD4, CD8, and CD20 in 206 thyroid carcinomas, 105 benign nodules, and 18 normal tissues. Also, TP53 was sequenced in 78 out of 164 patients with papillary thyroid carcinoma. Results. P53 expression was observed more frequently in malignant than in benign lesions (P < 0.0001) and helped discriminate follicular patterned lesions. In addition, p53 was more frequent in smaller (P = 0.0015), unique tumors (P = 0.0286), with thyroiditis (P = 0.0486) and without metastasis at diagnosis (P = 0.0201). TAM was more frequent in P53 negative tumors (P = 0.002). Infiltration of CD8+ TIL was found in 61.7% of P53 positive and 25.6% of P53 negative DTC (P < 0.001). Conclusions. We suggest that p53 and CD8+ TIL immune profile analysis might be useful in DTC.

Prognostic utility of basaloid differentiation in oropharyngeal cancer

BackgroundHuman papillomavirus (HPV) is recognized as the key risk factor for a distinct subset of oropharyngeal squamous cell carcinoma. P16 is a reliable, sensitive surrogate marker for HPV and confers a positive prognostic advantage. Basaloid differentiation on hematoxylin and eosin (H&E) staining is anecdotally noted by some pathologists to be associated with p16 positivity. This association, however, has not been adequately quantified in the literature, nor has the prognostic implications of basaloid differentiation been described.Objectives1) To correlate the H&E staining feature of basaloid differentiation with p16 positivity in oropharyngeal cancer. 2) To investigate the prognostic utility of basaloid differentiation in oropharyngeal cancer survival.MethodsRetrospective cross-sectional study of all patients diagnosed with and treated for oropharyngeal cancer at a single tertiary cancer center from 2002 to 2009. Tissue microarrays (TMAs) were generated from 208 oropharyngeal tumor specimens stained with H&E and immunohistochemical markers. These oropharyngeal TMAs were utilized in several previous publications. Samples were scored for basaloid differentiation by a pathologist blinded to the p16 result. A multivariate survival analysis with Cox-regression and Kaplan-Meier survival analysis was performed.ResultsIn the 208 samples, basaloid differentiation correlated with p16 positivity (Spearman’s rho 0.435). Basaloid differentiation and p16 positivity were both independent predictors of improved survival. The 5 year disease specific survival (DSS) was 73% for p16 positive tumors and 35% for p16 negative tumors (p < 0.001). Similarly, the 5 year DSS of basaloid differentiated tumors was 74% compared to 41% for non-basaloid tumors (p = 0.001). Patients with p16 positive and basaloid differentiated tumors had the best survival outcomes with a 5 year DSS of 80%.ConclusionsBasaloid differentiation is a feature on H&E which correlates with p16 positivity and is a simple, inexpensive, independent, positive prognostic indicator of comparable magnitude to p16 status. Due to the added prognostic value of basaloid differentiation, this feature should be routinely reported by qualified pathologists.

Biopsy Specimens Obtained 7 Days After Starting Chemoradiotherapy (CRT) Provide Reliable Predictors of Response to CRT for Rectal Cancer

Preoperative chemoradiation therapy (CRT) significantly decreases local recurrence in locally advanced rectal cancer. Various biomarkers in biopsy specimens obtained before CRT have been proposed as predictors of response. However, reliable biomarkers remain to be established. The study group comprised 101 consecutive patients with locally advanced rectal cancer who received preoperative CRT with oral uracil/tegafur (UFT) or S-1. We evaluated histologic findings on hematoxylin and eosin (H&E) staining and immunohistochemical expressions of Ki67, p53, p21, and apoptosis in biopsy specimens obtained before CRT and 7 days after starting CRT. These findings were contrasted with the histologic response and the degree of tumor shrinkage. In biopsy specimens obtained before CRT, histologic marked regression according to the Japanese Classification of Colorectal Carcinoma (JCCC) criteria and the degree of tumor shrinkage on barium enema examination (BE) were significantly greater in patients with p21-positive tumors than in those with p21-negative tumors (P=.04 and P<.01, respectively). In biopsy specimens obtained 7 days after starting CRT, pathologic complete response, histologic marked regression according to both the tumor regression criteria and JCCC criteria, and T downstaging were significantly greater in patients with apoptosis-positive and p21-positive tumors than in those with apoptosis-negative (P<.01, P=.02, P=.01, and P<.01, respectively) or p21-negative tumors (P=.03, P<.01, P<.01, and P=.02, respectively). The degree of tumor shrinkage on both BE as well as MRI was significantly greater in patients with apoptosis-positive and with p21-positive tumors than in those with apoptosis-negative or p21-negative tumors, respectively. Histologic changes in H&E-stained biopsy specimens 7 days after starting CRT significantly correlated with pathologic complete response and marked regression on both JCCC and tumor regression criteria, as well as with tumor shrinkage on BE and MRI (P<.01, P<.01, P<.01, P<.01, and P=.03, respectively). Immunohistochemical expressions of p21 and apoptosis together with histologic changes on H&E-stained biopsy specimens obtained 7 days after starting CRT are strong predictors of the response to CRT.

Nuclear epidermal growth factor receptor and p16 expression in head and neck squamous cell carcinoma

Epidermal growth factor receptor (EGFR) and p16 (a surrogate marker of human papillomavirus [HPV] infection) expression are strong prognostic factors in patients with head and neck squamous cell carcinoma (HNSCC). We examined expression levels of total and nuclear EGFR as well as p16 status based on evidence that nuclear EGFR may have a role in DNA damage repair. An HPV-negative (SQ20B) and an HPV-positive (UMSCC47) HNSCC cell line were examined for EGFR and γH2AX expression. A tissue microarray containing 123 cores obtained from 101 HNSCC tumors was analyzed for EGFR expression by automated quantitative analysis and p16 expression by immunohistochemical staining, and these results were correlated with available clinical data. SQ20B had higher EGFR expression than UMSCC47. Nuclear localization of EGFR on activation with transforming growth factor-α was observed in SQ20B, but not in UMSCC47. SQ20B also had increased γH2AX foci compared to UMSCC47, suggesting that SQ20B has more DNA damage compared to UMSCC47. Total and nuclear EGFR was reliably obtained from 80 of 101 patients. p16 levels were determined in 87 of 101 patients. p16 levels were strongly associated with the oropharyngeal subsite and poorly differentiated histology. Expression of total and nuclear EGFR was higher in p16-negative tumors compared to p16-positive tumors (Wilcoxon rank test, P = .038 and P = .014, respectively). Further studies are required to determine a mechanistic link between these two prognostic factors and the significance of EGFR localization to nucleus in DNA damage repair pathway activation.

575P - Biopsy Specimens Obtained 7 Days After Starting Chemoradiotherapy (CRT) Provides Reliable Predictors of Response to CRT for Rectal Cancer

ABSTRACT Background Preoperative CRT is a standard treatment for locally advanced rectal cancer (LARC). The histologic response to CRT or the downstaging has been reported to be closely related to oncologic outcomes. Various biomarkers in biopsy specimens obtained before CRT have been investigated as predictors of response, however, reliable predictive biomarkers remain to be established. Methods The study group comprised 101 consecutive patients with LARC who received preoperative CRT of 45Gy with oral uracil/tegafur (UFT) or S-1. We evaluated histologic findings on H-E staining and immunohistochemical expressions of Ki67, p53, p21, and apoptosis in biopsy specimens obtained before CRT and 7 days after starting CRT. These findings were contrasted with the rate of histologic marked regression and the degree of tumor shrinkage. Results In biopsy specimens obtained before CRT, the degree of tumor shrinkage on barium enema (BE) were significantly greater in patients with p21-positive tumors (52 ± 11%) than in those with p21-negative tumors (45 ± 16%) (p Conclusions Immunohistochemical expressions of p21 and apoptosis together with histologic changes on H-E-stained biopsy specimens obtained 7 days after starting CRT are strong predictors of response to CRT. Disclosure All authors have declared no conflicts of interest.

P16 Not a Prognostic Marker for Hypopharyngeal Squamous Cell Carcinoma

To investigate the prognostic significance of p16 in patients with hypopharyngeal squamous cell carcinoma (HPSCC) and to evaluate the relationship between p16 and human papillomavirus (HPV). Unlike in oropharyngeal SCC (OPSCC), the prognostic significance of p16 in HPSCC and its association with HPV is unclear. Retrospective medical chart review. University tertiary referral center. A total of 27 patients with HPSCC treated with definitive radiation therapy between 2002 and 2011 whose tissue was available for immunohistochemical analysis. Twenty-two patients were treated with chemoradiation, and 5 with radiation alone. All tumor biopsy specimens were analyzed for p16 and, when sufficient tissue was available, for HPV DNA. Overall survival (OS), locoregional control (LRC), disease-free survival (DFS), and laryngoesophageal dysfunction-free survival (LEDFS) were analyzed according to p16 status. Findings for p16 were positive in 9 tumors and negative in 18 tumors. Median follow-up was 29.3 months. There was no significant difference in OS, LRC, DFS, or LEDFS for patients with p16-positive vs p16-negative tumors. Only 1 of the 19 tumors tested for HPV was found to be HPV positive. When used as a test for HPV, p16 had a positive predictive value of 17%. In contrast to OPSCC, p16 expression in patients with HPSCC had a low positive predictive value for HPV and did not predict improved OS, LRC, DFS, or LEDFS. Thus, for HPSCC, p16 is not a prognostic biomarker. Caution must be taken when extrapolating the prognostic significance of p16 in patients with OPSCC to patients with head and neck SCC of other subsites.

Prognostic Significance of p16 Expression in Advanced Cervical Cancer Treated With Definitive Radiotherapy

The purpose of this study was to evaluate the prognostic significance of p16 immunohistochemistry (IHC) in patients with advanced cervical cancer treated with radiation therapy. This was a retrospective study of 126 patients with International Federation of Gynecology and Obstetrics Stages Ib1-IVb cervical cancer treated with radiation. Concurrent cisplatin chemotherapy was given to 108 patients. A tissue microarray (TMA) was constructed from the paraffin-embedded diagnostic biopsy specimens. Immunoperoxidase staining was performed on the TMA and a p16 monoclonal antibody was utilized. IHC p16 extent was evaluated and scored in quartiles: 0 = no staining, 1 = 1-25% of cells staining, 2 = 26 to 50%, 3 = 51 to 75%, and 4 = 76 to 100%. The p16 IHC score was 4 in 115 cases, 3 in 1, 2 in 3 and 0 in 7. There was no relationship between p16 score and tumor histology. Patients with p16-negative tumors were older (mean age at diagnosis 65 vs. 52 years for p16-positive tumors; p = 0.01). The 5-year cause-specific survivals were 33% for p16-negative cases (score = 0) compared with 63% for p16-positive cases (scores 1, 2, 3 or 4; p = 0.07). The 5-year recurrence-free survivals were 34% for those who were p16-negative vs. 57% for those who were p16-positive (p = 0.09). In addition, patients with p16-positive tumors (score > 0) were more likely to be complete metabolic responders as assessed by the 3-month posttherapy 18 [F]-fluorodeoxyglucose positron emission tomography (FDG-PET)/computed tomograph compared with patients with p16-negative tumors (p = 0.03). p16 expression is predictive of improved survival outcome after chemoradiation therapy for advanced-stage invasive cervical carcinoma. Further testing will be needed to evaluate p16-negative cervical tumors.

The apoptosis regulators p53, bax and PUMA: Relationship and impact on outcome in early stage (FIGO I-II) ovarian carcinoma after post-surgical taxane-based treatmen

The objective of this study was to evaluate the prognostic effect of the apoptosis regulators p53, bax and PUMA for recurrent disease and disease-free survival (DFS) in a series of 105 patients in FIGO-stagesI-II with epithelial ovarian cancer, all treated with post-surgical platinum-taxane chemotherapy. For the detection of positivity of the biological markers p53, bax and PUMA the techniques of tissue microarrays and immunohistochemistry (IHC) were used. In tumors the frequency of p53 positivity was 24%, that of bax positivity was 83%, and strong positivity was found for PUMA (43%). The bax status was related to tumor grade (P=0.029). Positive staining for bax was related to strong positivity of PUMA in the tumors (P=0.004). The p53, bax or PUMA status alone or concomitant (p53 bax, p53 PUMA and bax PUMA) were not related to age, histopathological subtype, serous/non-serous tumors or type of the staging procedure at primary surgery. In survival analysis p53-positive tumors (P=0.014) and concomitant p53-positive and weak PUMA-positive tumors (P=0.015) were significantly correlated with shorter DFS. Concomitant p53-negative and bax-positive tumors were significantly correlated with longer survival (P=0.019). FIGO-stage (OR=6.0) and p53 status (OR=4.1) were predictive factors for tumor recurrence in logistic regression analysis and independent prognostic factors (HR=2.4 for both) in multivariate Cox regression analysis. In a separate Cox multivariate regression analysis the p53 bax status (HR=2.2) was an independent prognostic factor for DFS. The p53 PUMA status (HR=0.4) was not an independent prognostic factor, however, a borderline significance (P=0.07) was noted. Our results indicate that FIGO stage and p53 status alone were independent predictive factors for recurrence and prognostic factors for survival. Furthermore, p53 bax status was an independent prognostic factor for survival in this study.

Wip1 promotes RUNX2-dependent apoptosis in p53-negative tumors and protects normal tissues during treatment with anticancer agents

The inactivation of the p53 tumor suppressor pathway in many cancers often increases their resistance to anticancer therapy. Here we show that a previously proposed strategy directed to Wip1 inhibition could be ineffective in tumors lacking p53. On the contrary, Wip1 overexpression sensitized these tumors to chemotherapeutic agents. This effect was mediated through interaction between Wip1 and RUNX2 that resulted, in response to anticancer treatment, in RUNX2-dependent transcriptional induction of the proapoptotic Bax protein. The potentiating effects of Wip1 overexpression on chemotherapeutic agents were directed only to tumor cells lacking p53. The overexpression of Wip1 in normal tissues provided protection from cisplatin-induced apoptosis through decreased strength of upstream signaling to p53. Thus, Wip1 phosphatase promotes apoptosis in p53-negative tumors and protects normal tissues during treatment with anticancer agents.

A clinical, pathologic, and molecular study of p53 and murine double minute 2 in penile carcinogenesis and its relation to prognosis

Penile carcinoma constitutes up to 16% of male malignancies in developing countries. Changes in the p53 and murine double minute 2 pathway are important events in various cancers. Associate alterations in murine double minute 2 and p53 expression were evaluated by molecular techniques, with the clinical data of 297 cases of penile carcinoma. Automated immunohistochemistry was performed for murine double minute 2 and p53 using the primary antibodies SPM14 and DO7, respectively. Fluorescent in situ hybridization was performed using the probes murine double minute 2 at 12q15 and TP53 at 17p13.1. Slides were digitalized, and bright-field and fluorescent images were analyzed. TP53 was sequenced in 16 cases. The expression of p53 was higher in poorly differentiated, infiltrative border, corpus spongiosum, corpora cavernosa, and invasive urethral carcinomas. Patients who died of disease also expressed higher levels of p53. p53-negative tumors were associated with higher overall survival. Murine double minute 2 showed no difference of expression in any group of tumors, no correlation with p53 expression. No alterations in genes or chromosomes were observed. Mutations in TP53 were observed in 4 of 16 cases: p.T170M, p.L252P, p.C176Y, and the novel c.803_810del8; these changes correlated with p53 expression by immunohistochemistry. Murine double minute 2 is not useful in the prognosis of penile carcinoma by immunohistochemistry. Additional studies on the transcriptional, posttranscriptional, and epigenetic aspects are necessary to understand the interactions between p53 and murine double minute 2 because we did not observe any numeric alterations by fluorescent in situ hybridization. Examining p53 is helpful in identifying patients with more aggressive tumors and may be crucial in selecting the most suitable surgical procedure.