Maria Gonidi

Apoptotic and inflammation markers in oral mucositis in head and neck cancer patients receiving radiotherapy : preliminary report

Goal of workThe aim of this study was to investigate the expression of pro-apoptotic protein p53 and anti-apoptotic proteins BCl-2 and MCl-1, as well as the expression of pro-inflammatory cytokines tumor necrosis factor (TNF) and interleukin-1beta (IL-1β) in patients developing mucositis during radiotherapy for head and neck cancer.Materials and methodsThirty-five patients receiving radiotherapy for head/neck cancer were included in this study. Patients were examined before radiotherapy. Oral mucositis was recorded weekly during radiotherapy. Cytologic smears from the oral cavity were taken with a brush. Immunocytochemical staining was performed by the use of p53, BCl-2, MCl-1 TNF and IL-1β monoclonal antibodies.Main resultsP53 was expressed in 1 of 15 smears before the initiation of radiotherapy (6.5%) compared to 3 of 7 smears from patients with grade III mucositis (43%) during radiotherapy. BCl-2 was expressed in 15 of 15 smears before radiotherapy (100%) and in three of seven patients with grade III mucositis (43%) during radiotherapy. MCl-1 was expressed in 10 of 14 samples before radiotherapy (71.5%) and in two of seven patients with grade III (28.5%) mucositis during radiotherapy. TNF was expressed in 9 of 14 patients before radiotherapy (64%) and in six of seven patients with grade III mucositis during radiotherapy (86%). IL-1β was detected in 7 of 14 patients before radiotherapy (50%) compared to 6 of 7 patients with grade III mucositis during radiotherapy (86%).ConclusionOur preliminary results indicate an induction of apoptosis and inflammation in the oral mucosa in patients developing mucositis during radiotherapy for head/neck cancer.

CD24 expression has a prognostic impact in breast carcinoma.

We investigated the prognostic significance of BAG-1 and CD24 in invasive breast carcinomas. Seventy cases of invasive breast carcinoma were studied immunocytochemically for the expression of BAG-1 and CD24. The results were correlated with several prognostic parameters, including 5-year survival. Univariate analysis showed a significant correlation of BAG-1 and CD24 overall positive staining with several adverse prognostic parameters, such as increased stage (p<0.0001), tumor grade 3 (p=0.016 and p=0.02, respectively), positive lymph nodes (p<0.0001), and increased tumor size (p<0.0001). Similar results were found for BAG-1 nuclear staining, as well as for positive cytoplasmic CD24 expression. Both of our markers studied had a significant, negative effect on survival. Multivariate analysis further revealed an independent prognostic impact for CD24 overall staining. The results of our study showed that overall cytoplasmic and especially nuclear BAG-1 expression, as well as overall and cytoplasmic CD24 expression, correlates with adverse prognostic parameters. An independent prognostic value for overall CD24 staining was also demonstrated.

The significance of Survivin and Nectin-4 expression in the prognosis of breast carcinoma.

To investigate the prognostic significance of Survivin and Nectin-4 expression in breast carcinomas. Imprint smears were obtained from 140 breast carcinoma specimens and studied immunocytochemically for the expression of Survivin and Nectin-4. The results were correlated with several clinicopathological parameters, including five-year survival. Increased Survivin staining pattern correlated with increased grade (p < 0.0001), increased lymph node invasion (p < 0.0001), increased tumor size and reduced survival (p < 0.0001). Elevated Nectin-4 expression also correlated significantly with increased grade (p < 0.0001), increased tumor size (p < 0.0001) and reduced survival (p < 0.0001). In addition, Survivin and Nectin-4 staining patterns correlated strongly with one another (p < 0.0001). However, on multivariate analysis, neither Survivin nor Nectin-4 expression seemed to have an independent impact on survival in our study cases. The findings of our study suggest that increased expression of Survivin and Nectin-4 may indicate a worse prognosis in breast cancer patients. The exact implications of the expression of these markers in breast cancer prognosis and treatment remain to be clarified.

Mitochondrial UCP4 and bcl-2 expression in imprints of breast carcinomas: Relationship with DNA ploidy and classical prognostic factors

Mitochondria are the bioenergetic and metabolic centers of cells and play an important role in the regulation of cell death. The mitochondrial apoptosis pathway is controlled by the bcl-2 protein family. Overexpression of mitochondrial uncoupling protein 4 (UCP4) can promote proliferation and inhibit apoptosis and differentiation. Imprint smears obtained from 124 tumors were studied immunocytochemically, and results were correlated with prognostic markers. There were 112 ductal and 12 lobular carcinomas. The positivity of UCP4 was correlated with lymph node metastases (p=0.005), positive ER and PR expression (p<0.0001 for both), as well as positivity for p53 (p<0.0001) and Ki-67 (p<0.0001). Decreased expression of bcl-2 correlated with increased expression of UCP4 (p=0.001). Regarding DNA ploidy, UCP4 positivity was correlated with aneuploid tumors (p=0.002). Negative expression of bcl-2 was correlated with poorly differentiated carcinomas (p<0.0001), as well as with positive expression of p53 (p<0.0001) and Ki-67 (p<0.0001). Logistic regression revealed that ploidy and p53 expression had an impact on UCP4. These findings encourage future investigations regarding the potential role of UCPs not only into mechanisms underlying breast cancer, but also as a novel candidate to the design and development of more effective therapeutic strategies.

Enhancer of Zeste Homologue 2 Expression in Breast Carcinoma Smears in Relationship with p53, Ki-67 and Other Prognostic Parameters

Background: The polycomb group protein enhancer of zeste homologue 2 (EZH 2) has been reported as a marker of aggressive breast cancer. The aim of this study was to investigate the correlation between the expression of EZH 2 with p53 and Ki-67 expression and other clinicopathological parameters in primary breast carcinomas in order to determine the role of the above marker as a prognosticator of tumor aggressiveness and patient outcome. Patients and Methods: One hundred primary operable breast cancer patients were investigated in order to identify the expression of EZH 2, Ki-67 and p53 in imprint smears immunocytochemically. The prevalence of expression of these markers was then correlated with clinicopathological parameters. Follow-up was available for all patients. Results: EZH 2 was expressed in 64% of the cases and correlated with higher levels of p53 (relative risk = 3.00, p < 0.0001) and Ki-67 (relative risk = 3.25, p < 0.0001). Malignant cells showed immunoreactivity for all markers in the nucleus. Univariate analysis revealed a strong association between EZH 2 protein expression and tumor grade and size, lymph node metastasis, and HER-2 and estrogen and progesterone receptor status. Multivariable statistical analysis revealed that lymph node metastasis was the main predictor for EZH 2 expression. Decreased patient survival was also significantly associated with EZH 2 expression (p < 0.0001). Conclusions: EZH 2 expression may be a marker of poor prognosis in breast carcinoma patients and has been suggested as a candidate for targeted therapy.

Moc-31, fibronectin and CEA in the differential diagnosis of malignant effusions: an immunocytochemical study.

In discriminating benign and malignant origins of cytologically suspicious effusion smears a panel of antibodies against carcinoembryonic antigen (CEA), Fibronectin (F) and MOC-31 was used with immunocytochemical techniques. One hundred and thirty seven effusions were studied of which 107 had a malignant and 30 a benign aetiology as determined by clinical and histological examination. Cytologically 24 were diagnosed as benign, 97 as malignant and 14 as suspicious. Staining for F was positive in all effusions of benign and 3 of malignant origin. MOC-31 was positive in 95 (88.8%) of effusions of malignant origin but none of benign origin. Positive CEA was observed in 43% of effusions of malignant origin and in 10 of benign origin. The combination of MOC-31 positivity measured the sensitivity and specificity of the cytological examination in cases where the cytological examination result was suspicious as did F positivity improve the sensitivity for a benign origin of the effusion. Positivity or negativity for CEA is less valuable than the other parameters.

Primary malignant melanoma of the conjunctiva of the upper eyelid : A case report

BACKGROUND Malignant melanoma of the conjunctiva is rare. The nomenclature and clinical and pathologic features of cutaneous and conjunctival melanomas are different. CASE A 62-year-old male presented with a history of slight bleeding of the upper conjunctiva for the previous six months. On clinical examination the ophthalmologist observed a smooth, partly nodular, pigmented lesion on the conjunctiva under the left eyelid, 1.5 cm in diameter. Fine needle aspiration (FNA) biopsy of the mass showed tumor cells dispersed as single cells with eccentric, round nuclei; coarsely granular chromatin; prominent nucleoli; and dense cytoplasm with occasional brownish pigmentation as well as small aggregates of spindle-shaped neoplastic cells with hyperchromatic nuclei and no cytoplasmic pigment. CONCLUSION FNA cytology is a simple and efficient method of making the diagnosis of malignant melanoma in conjunctival masses. Careful correlation with the clinical history and histologic findings is often necessary for confirmation of the diagnosis.

Prognostic value of COX‐2, P53, and EZH‐2 evaluated by quantitative image analysis in premalignant and malignant breast lesions

Cytological differential diagnosis of atypical hyperplasia and well differentiated breast carcinoma may be challenging, because sometimes there is an overlap between the cytomorphological features of these lesions. The aim of the study was to investigate COX‐2, EZH‐2, p53 expression in carcinomas and the gray zone of breast cytology categories of atypical hyperplastic lesions with regard to biological behavior of the tumor.

Immunocytochemical expression of a panel of markers in pleural effusions from patients with primary lung adenocarcinoma

INTRODUCTION This studys principal objective was to evaluate the critical role of the application of immunocytochemistry to a novel panel of diagnostic markers for the accurate detection of the source of malignancies in pleural effusions of lung adenocarcinoma. MATERIALS AND METHODS In 40 effusion smears from lung adenocarcinoma, the expression of the E-cadherin, a-catenin, Thyroid Transcription Factor (TTF-1), Epidermal Growth Factor Receptor (EGFR), p53, caspase 9 and 3, Bax and Bcl-2 was examined by immunocytochemistry. RESULTS All cases showed positive immunoreactivity of tumour cells to caspase 3 (42,5%), caspase 9 (40%), Bcl-2 (30%), Bax (40%), p53 (55%), E-cadherin (82,5%), a-catenin (80%), TTF-1 (87,5%) and EGFR (62,5%). The Pearsons x2 analysis demonstrated a highly significant correlation to each of the other marker when analysed separately. Caspase 3 expression was correlated significantly with caspase 9 (p<0.0001), Bax (p=0.002), Bcl-2(p=0.014) and p53 (p=0.011). Caspase 9 was correlated with Bax (p=0.005) and p53 (p=0.047), p53 correlated with E-cadherin (p=0.011), a-catenin(p=0.011), EGFR (p<0.0001) and Bax (p=0.032). Correlation was also observed between Bcl-2 and Bax expression (p<0.0001), E-cadherin and a-catenin expression (p<0.0001) and a-catenin and TTF-1 expression (p=0.002). CONCLUSIONS The use of a panel of biomarkers can be of great value in determining effusion immunoprofile in patients with lung adenocarcinoma for clinical application.

13P Expression of Notch 1 and Notch 3 proteins in imprints of NSCLC: Correlation with prognostic factors

Background: Mutations in the exon 14 of the MET gene affecting RNA splice acceptor and donor sites, which lead to exon skipping, have been recently associated to responsiveness to crizotinib. Up to date, patient selection for Met inhibitors has been made according to MET amplification/high polysomy and/or protein overexpression. We have investigated the prevalence of MET exon 14 mutations and their correlation with gene copy number alterations (CNAs) in NSCLC patients. Methods: We tested 132 lung adenocarcinomas and adenosquamous carcinoma samples for mutations in MET exon 14 by Sanger sequencing and for MET gene CNAs by FISH (Abbott Molecular). We collected clinical data together with EGFR and KRASmutational status and ALK, ROS1 and RET rearrangements. Results: MET alterations were found in 23 patients (17.5%): 3 exon 14 skipping mutations (2.3%), 8 gene amplifications (6.1%), and 12 high polysomy cases (9.1%). Among the 11 MET altered cases (3 mutated and 8 amplified): 8 patients were male, with a median age of 64.5 years (range: 46–91), current or former smokers (50 pack/years, range: 30–80), and all were diagnosed in an advanced stage disease (III and IV). None of the 3 mutated cases had concurrent MET CNAs. One MET mutated case presented also a KRAS p.G12C substitution while the other two were wild type for EGFR (exons 18 to 21) and KRAS (exon 2). Interestingly, patients with MET alterations have a shorter overall survival (p < 0.001). Conclusions: We have identified alterations of the MET gene (mutations in exon 14 and CNAs) in 8.4% of NSCLC patients. Our data support the usefulness of prospective screening of MET exon 14 mutations and gene amplifications due to their prognostic and predictive role. These mutations define a new subset of NSCLC patients that should be analyzed independently of the MET gene copy number status. Legal entity responsible for the study: Institut Hospital del Mar d’Investigacions Mèdiques (IMIM), Barcelona, Spain Funding: Institut Hospital del Mar d’Investigacions Mèdiques (IMIM), Barcelona, Spain Disclosure: All authors have declared no conflicts of interest.