Relapse in dermatofibrosarcoma protuberans: A histological and molecular analysis.

BACKGROUND: Dermatofibrosarcoma protuberans (DFSP) is a rare low grade tumor with a locally aggressive behavior and low metastatic potential. OBJECTIVES: To evaluate the factors that are associated with relapse in DFSP. Methods Retrospective analysis of medical records from 61 patients with dermatofibrosarcoma. Fluorescence in situ hybridization was used to detect translocations. RESULTS: Of 61 patients, 6 experienced a relapse. No patient with resection margins greater than 3 cm had a recurrence. One relapse was observed in a patient treated with at least 2 cm margins and 4 relapses occurred in 16 patients whose margins were below 2 cm (P = 0.018). The frequency of translocations was 77.8%. The recurrence rate was lower in patients with translocation, but this difference was not significant. Immunohistochemical markers did not correlate with recurrence rates, but greater FasL expression was associated with recurrence in patients with margins smaller than 3 cm. CONCLUSIONS: Surgical margins smaller than than 2 cm are related to higher recurrences in dermatofibrosarcomas. In this analysis a 2 cm margin was acceptable for treatment. Between all the immunohistochemical markers analyzed, only FasL was associated with a higher recurrence rate in patients with margins smaller than 3 cm.

Multi-task network for automated analysis of high-resolution endomicroscopy images to detect cervical precancer and cancer.

Cervical cancer is a public health emergency in low- and middle-income countries where resource limitations hamper standard-of-care prevention strategies. The high-resolution endomicroscope (HRME) is a low-cost, point-of-care device with which care providers can image the nuclear morphology of cervical lesions. Here, we propose a deep learning framework to diagnose cervical intraepithelial neoplasia grade 2 or more severe from HRME images. The proposed multi-task convolutional neural network uses nuclear segmentation to learn a diagnostically relevant representation. Nuclear segmentation was trained via proxy labels to circumvent the need for expensive, manually annotated nuclear masks. A dataset of images from over 1600 patients was used to train, validate, and test our algorithm; data from 20% of patients were reserved for testing. An external evaluation set with images from 508 patients was used to further validate our findings. The proposed method consistently outperformed other state-of-the art architectures achieving a test per patient area under the receiver operating characteristic curve (AUC-ROC) of 0.87. Performance was comparable to expert colposcopy with a test sensitivity and specificity of 0.94 (p = 0.3) and 0.58 (p = 1.0), respectively. Patients with recurrent human papillomavirus (HPV) infections are at a higher risk of developing cervical cancer. Thus, we sought to incorporate HPV DNA test results as a feature to inform prediction. We found that incorporating patient HPV status improved test specificity to 0.71 at a sensitivity of 0.94.

MicroRNA Biomarkers of High-Grade Cervical Intraepithelial Neoplasia in Liquid Biopsy.

New prevention strategies are needed to detect cervical intraepithelial neoplasia (CIN). The microRNA expression analysis has already been reported as molecular biomarkers in the early detection of cervical cancer (CC) through minimally invasive samples, such as liquid biopsy, obtained through collection using liquid-based cytology (LBC). In this study, we aimed to identify molecular signatures of microRNAs in cervical precursor lesions from LBC cervical and the molecular pathways potentially associated with the CC progression. We analyzed 31 LBC cervical samples from women who underwent colposcopy. These samples were divided into two groups: the first group was composed of samples without precursor lesions of CC, considering the control group, referred to as healthy female subjects (HFS; n = 11). The second group corresponded to women diagnosed with cervical interepithelial neoplasia grade 3 (CIN 3; n = 20). We performed microRNA and gene expression profiling using the nCounter® miRNA Expression Assays (NanoString Technology) and PanCancer Pathways (NanoString Technology), respectively. A microRNA target prediction was performed by mirDIP, and molecular pathway interaction was constructed using Cytoscape. Bidirectional in silico analyses and Pearson's correlation were performed for associated the relation between genes, and miRNAs differentially expressed related cervical cancer progression were performed. We found that the expression of nine microRNAs was significantly higher, two were downregulated (miR-381-3p and miR-4531), and seven miRNAs were upregulated (miR-205-5p, miR-130a-3p, miR-3136-3p, miR-128-2-5p, let-7f-5p, miR-202-3p, and miR-323a-5p) in CIN 3 (fold change ≥ 2 and p ≤ 0.05). The miRNA expression patterns were independent of hr-HPV infection. We identified four miRNAs (miR-205-5p, miR-130a-3p, miR-4531, and miR-381-3p) that could be used as biomarkers for CIN 3 in LBC samples through multiple logistic regression analyses. We found 16 genes differentially expressed between CIN 3 and HSF samples (fold change ≥ 2 and p ≤ 0.05). We found the correlation between miR-130a-3p and CCND1(R = -0.52; p = 0.0029), miR-205-5p and EGFR (R = 0.53; p = 0.0021), and miR-4531 and SMAD2 (R = -0.54; p = 0.0016). In addition, we demonstrated the most significant pathways of the targets associated with cervical cancer progression (FDR-corrected p < 0.001). This study demonstrated that miRNA biomarkers may distinguish healthy cervix and CIN 3 and regulate important molecular pathways of carcinogenesis.

Topical Imiquimod for the Treatment of High-Grade Squamous Intraepithelial Lesions of the Cervix: A Randomized Controlled Trial.

OBJECTIVE: To evaluate the histologic response rate of high-grade squamous intraepithelial lesions (HSIL) of the cervix after topical application of 5% imiquimod cream. METHODS: In this phase II trial, women with cervical HSIL (cervical intraepithelial neoplasia [CIN] 2-3) were randomly assigned to 250 mg of 5% imiquimod cream applied to the cervix weekly for 12 weeks, followed by loop electrosurgical excision procedure (LEEP) without preceding treatment. The sample size was calculated based on the HSIL regression rates previously reported by Grimm et al. The primary outcome was rate of histologic regression (to CIN 1 or less) in LEEP specimens. Prespecified secondary endpoints included surgical margin status and adverse events. Outcomes were stratified by human papillomavirus type and lesion grade (CIN 2 or CIN 3). Results were reported according to per protocol (PP) and intention-to-treat (ITT) analyses. RESULTS: Ninety women were enrolled: 49 in the experimental group and 41 in the control group. In the PP population, histologic regression was observed in 23 of 38 participants (61%) in the experimental group compared with 9 of 40 (23%) in the control group (P=.001). Surgical margins were negative for HSIL in 36 of 38 participants (95%) in the experimental group and 28 of 40 (70%) in the control group (P=.004). In the ITT population, rates of histologic regression also were significantly higher in the experimental group. Rates of adverse events in the experimental group were 74% (28/38) in the PP population and 78% (35/45) in the ITT population. Adverse events were mild, with abdominal pain being the most common. Three patients in the experimental group had grade 2 adverse events, including vaginal ulcer, vaginal pruritus with local edema, and moderate pelvic pain. CONCLUSION: Weekly topical treatment with imiquimod is effective in promoting regression of cervical HSIL. CLINICAL TRIAL REGISTRATION: ClinicalTrials.gov, NCT03233412.

Evaluation of gene amplification and protein expression of HER-2/neu in esophageal squamous cell carcinoma using Fluorescence in situ Hybridization (FISH) and immunohistochemistry.

BACKGROUND: Esophageal squamous cell carcinoma (ESCC) is the sixth most frequent neoplasia in Brazil. It is usually associated with a poor prognosis because it is often at an advanced stage when diagnosed and there is a high frequency of lymph node metastases. It is important to know what prognostic factors can facilitate diagnosis, optimize therapeutic decisions, and improve the survival of these patients. A member of the epidermal growth factor receptor (EGFR) family, c-erbB-2, has received much attention because of its therapeutic implications; however, few studies involving fluorescence in situ hybridization (FISH) analysis of HER-2/neu gene amplification and protein expression in ESCC have been conducted. The aim of this study was to verify the presence of HER-2/neu gene amplification using FISH, and to correlate the results with immunohistochemical expression and clinical-pathological findings. METHODS: One hundred and ninety-nine ESCC cases were evaluated using the Tissue Microarray (TMA) technique. A polyclonal antibody against c-erbB-2 was used for immunohistochemistry. Analyses were based on the membrane staining pattern. The results were classified according to the Herceptest criteria (DAKO): negative (0/1+), potential positive (2+) and positive (3+). The FISH reactions were performed according to the FISH HER2 PharmDx (DAKO) protocol. In each case, 100 tumor nuclei were evaluated. Cases showing a gene/CEN17 fluorescence ratio > or = 2 were considered positive for gene amplification. RESULTS: The c-erbB-2 expression was negative in 117/185 cases (63.2%) and positive in 68 (36.8%), of which 56 (30.3%) were 2+ and 12 (6.5%) were 3+. No significant associations were found among protein expression, clinicopathological data and overall survival. Among the 47 cases analyzed, 38 (80.9%) showed no gene amplification while 9 (19.1%) showed amplification, as demonstrated by FISH. Cases that were negative (0/1+) and potential positive (2+) for c-erbB-2 expression by immunohistochemistry showed no gene amplification. However, all cases with gene amplification were positive (3+) by immunohistochemistry. According to univariate analysis, there was a significant difference (p = 0.003) in survival rates when cases with and without HER-2/neu amplification were compared. CONCLUSION: Our data demonstrate the correspondence between gene amplification and protein expression of HER-2/neu. Gene amplification is an indicator of poor prognosis in ESCC.

Comparison of biological behavior between early-stage adenocarcinoma and squamous cell carcinoma of the uterine cervix.

OBJECTIVES: (1) To compare the anatomopathological variables and recurrence rates in patients with early-stage adenocarcinoma (AC) and squamous cell carcinoma (SCC) of the uterine cervix; (2) to identify the independent risk factors for recurrence. STUDY DESIGN: This historical cohort study assessed 238 patients with carcinoma of the uterine cervix (IB and IIA), who underwent radical hysterectomy with pelvic lymph node dissection between 1980 and 1999. Comparison of category variables between the two histological types was carried out using the Pearson's chi(2)-test or Fisher exact test. Disease-free survival rates for AC and SCC were calculated using the Kaplan-Meier method and the curves were compared using the log-rank test. The Cox proportional hazards model was used to identify the independent risk factors for recurrence. RESULTS: There were 35 cases of AC (14.7%) and 203 of SCC (85.3%). AC presented lower histological grade than did SCC (grade 1: 68.6% versus 9.4%; p<0.001), lower rate of lymphovascular space involvement (25.7% versus 53.7%; p=0.002), lower rate of invasion into the middle or deep thirds of the uterine cervix (40.0% versus 80.8%; p<0.001) and lower rate of lymph node metastasis (2.9% versus 16.3%; p=0.036). Although the recurrence rate was lower for AC than for SCC (11.4% versus 15.8%), this difference was not statistically significant (p=0.509). Multivariate analysis identified three independent risk factors for recurrence: presence of metastases in the pelvic lymph nodes, invasion of the deep third of the uterine cervix and absence of or slight inflammatory reaction in the cervix. When these variables were adjusted for the histological type and radiotherapy status, they remained in the model as independent risk factors. CONCLUSION: The AC group showed less aggressive histological behavior than did the SCC group, but no difference in the disease-free survival rates was noted.

High-resolution microendoscopy: a point-of-care diagnostic for cervical dysplasia in low-resource settings.

Cervical cancer is the third leading cause of cancer-related death among women in low-to-middle income countries. Pap testing and pathological services are difficult to implement under these settings. Alternative techniques for the diagnosis of cervical precancer in these settings are needed to reduce the burden of the disease. The objective of this study was to evaluate the diagnostic accuracy of a low-cost, high-resolution microendoscope imaging system in identifying precancerous lesions of the cervix in vivo. A retrospective study of 59 patients undergoing colposcopy for an abnormal Pap test was performed at Hospital de Câncer de Barretos in Brazil. All patients underwent colposcopy as per standard of care, and acetowhite lesions were recorded. High-resolution microendoscopy (HRME) images were obtained from one colposcopically normal region and from all lesions observed on colposcopy. Biopsies of abnormal areas were obtained and reviewed by three independent, blinded pathologists and compared with HRME findings. The mean nuclear area and the median nuclear eccentricity were calculated from HRME images acquired from each site. A diagnostic algorithm to distinguish histopathologically diagnosed cervical intraepithelial neoplasias of grade 2 or more severe lesions (high grade) from less severe lesions (low grade) was developed using these parameters. A test of trend was used to analyze the relationship between HRME positivity and severity of histopathogical diagnosis. Fisher's exact test was used to analyze differences in HRME positivity between high-grade and low-grade lesions. Evaluable images were obtained from 108 of 143 discrete sites. Of these, 71 sites were colposcopically normal or low grade according to histopathology and 37 were diagnosed as high grade on the basis of histopathology. Using the mean nuclear area and the median nuclear eccentricity, HRME images from 59 colposcopically abnormal sites were classified as high grade or low grade with 92% sensitivity and 77% specificity compared with histopathological findings. Increasing HRME positivity showed a significant trend with increasing severity of diagnosis (Ptrend<0.001). We found a strong association (P<0.001) between HRME positivity and a histopathological diagnosis of cervical intraepithelial neoplasia of grade 2 or higher. HRME demonstrated an accurate in-situ diagnosis of high-grade dysplasia. In low-resource settings in which colposcopy and histopathology services are severely limited or unavailable, HRME may provide a low-cost, accurate method for diagnosis of cervical precancer without the need for biopsy, allowing for a single 'screen-and-treat' approach.

Prognostic Significance of Apoptosis-related Markers in Patients With Soft-Tissue Sarcomas of Extremities.

Bcl-2 and Bax proteins are key regulators of apoptosis, a process that is deregulated in many human diseases, particularly cancer. Overexpression of antiapoptotic Bcl-2 protein is associated with drug resistance and poor clinical outcome in cancer patients, whereas the expression of proapoptotic Bax protein, commonly detected in soft-tissue sarcoma (STS), is often associated with chemiosensitivity in different tumors. Studies on the clinical implications of apoptosis-related markers Bcl-2 and Bax in STS are limited. In this study, immunohistochemistry for Bcl-2 and Bax was performed on tissue microarrays of 86 multiple types of adult STS of the extremities. Bcl-2 and Bax positive expression was detected in 25.9% and 66.7% of the sarcomas, respectively. Overexpression of both, Bcl-2 and Bax, was directly associated with histologic grade and clinical stage. A significant association between Bax and Bcl-2 expression was also observed (P=0.007). The 5-year overall survival for the group was 57%, and it was lower for cases that overexpressed Bcl-2 (47.6% vs. 58.3%) and Bax (50% vs. 66.7%), although not statistically significant. After multivariate analysis, only the high histologic grade appeared as an independent prognostic factor for the patients (P=0.043; HR=8.0; 95% CI, 1.1-60.1). In our study, Bcl-2 and Bax expression was significantly associated with histologic grade and clinical stage, which are classic factors of poor prognosis. We suggest the use of these proteins as potential prognostic markers in STS of extremities.

Randomized clinical trial comparing cold knife conization of the cervix with and without lateral hemostatic sutures.

OBJECTIVE: Compare blood loss during cold knife conization of the cervix with and without lateral hemostatic sutures in the cervical branches of the uterine arteries. DESIGN: Randomized clinical trial. SETTING: Hospital de Clínicas de Porto Alegre (HCPA). POPULATION: 102 patients that underwent cold knife conization. METHODS: Women that underwent cold knife conization of the cervix were randomized to undergo the procedure with or without lateral hemostatic sutures. PRIMARY OUTCOME MEASURE: blood loss measured in grams. SECONDARY OUTCOME MEASURES: operative time and postoperative intervention. Only the participants were blinded to group assignment. RESULTS: From March 2009 to August 2012, patients were randomly assigned to one of the study groups. There were no differences in amount of blood loss between patients that underwent the procedure with and without sutures (p = 0.39). Operative time was shorter in the group without suture (p = 0.020). There were no differences in intervention due to bleeding (p = 0.20). Blood loss was greater among menstruating women than for menopausal women (p = 0.011). There were no differences in amount of blood lost between smoking and nonsmoking patients (p = 0.082). CONCLUSIONS: Lateral hemostatic sutures do not affect the amount of intraoperative bleeding or the number of postoperative interventions. Their use is not necessary because they result in longer operative time, have a higher cost due to the use of suture material and pose the risk of ureter lesion in case the sutures are not placed at a lower position in the cervix. ClinicalTrials. gov identifier: NCT02184975.

The Accuracy of p16/Ki-67 and HPV Test in the Detection of CIN2/3 in Women Diagnosed with ASC-US or LSIL.

The objective of this study was to compare the accuracies of double staining for p16/Ki-67 and the molecular test for high-risk HPV (hr-HPV) to identify high-grade cervical intraepithelial neoplasia (CIN2/CIN3) in women with cervical cytology of atypical squamous cells of undetermined significance (ASC-US) and low-grade squamous intraepithelial lesion (LSIL). Data were collected from 201 women who underwent cervical cytology screening in the Barretos Cancer Hospital and their results were categorized as ASC-US (n=96) or LSIL (n=105). All patients underwent colposcopy with or without cervical biopsy for diagnosis of CIN2/CIN3. The hr-HPV test (Cobas 4800 test) and immunocytochemistry were performed to detect biomarkers p16/Ki-67 (CINtec PLUS test). Two samples (1 ASC-US/1 LSIL) were excluded from the analysis due to inconclusive results of the histologic examination. There were 8 cases of CIN2/CIN3 among 95 women with ASC-US (8.4%), and 23 cases of CIN2/CIN3 among 104 women with LSIL (22.1%). In the group of women with ASC-US, the sensitivity and specificity in diagnosing CIN2/CIN3 were 87.5% and 79.5% for the HPV test and 62.5% and 93.1% for p16/Ki-67. Among women with LSIL, the sensitivity and specificity in the diagnosis of CIN2/CIN3 were 87% and 34.7% for the HPV test and 69.6% and 75.3% for immunocytochemistry. Superior performance was observed for p16/Ki-67 double staining, especially among women under 30 for whom the test had an area under the ROC curve of 0.762 (p<0.001). Both p16/Ki-67 double staining and the hr-HPV DNA test had similar performance in predicting high-grade cervical intraepithelial neoplasia among women with ASC-US. The best performance was observed in women aged >30 years. In younger women (≤30 years) with LSIL, p16/Ki-67 had greater accuracy in identifying precursor lesions. Among women >30 years diagnosed with LSIL, the two methods showed similar performance.

Prognostic implications of altered human epidermal growth factor receptors (HERs) in gastric carcinomas: HER2 and HER3 are predictors of poor outcome.

PURPOSE: The human epidermal growth factor receptor (HER) family consists of four members: ErbB-1 (HER1), ErbB-2 (HER2), ErbB-3 (HER3), and ErbB-4 (HER4). These receptors activate numerous downstream pathways in response to extracellular ligands, regulating diverse processes that include differentiation, migration, proliferation, and survival. Alterations in these genes play a role in the development and progression of many human cancers. In gastric carcinomas (GCs), expression of HER1 and HER2 is thought to be a prognostic factor and target of novel biologic agents. The effect of HER3 or HER4 expression in GC has not been sufficiently studied. In this study, we explored the gene and protein expression of the HER family in GC to establish new potential prognostic factors. PATIENTS AND METHODS: Immunohistochemistry and fluorescence in situ hybridization were performed in 221 patients with GC using tissue microarray. Correlation between the expression or amplification of HER genes and the clinicopathologic parameters was statistically analyzed. RESULTS: Alterations of members of the HER family were significantly associated with the parameters involved in tumor progression, including depth of tumor invasion, involved lymph nodes, and tumor stage. In addition, HER2 amplification and HER3 expression were significantly related to worse survival. CONCLUSION: These results reveal that all members of the HER family are expressed in GC. Furthermore, expression of HER2 and HER3 is a significant predictor of poor survival in GC. Therefore, the development of HER-targeted agents and agents targeting downstream signaling pathways provides new possibilities in the treatment of GC.

Menopausal status: a possible predictive factor for recurrence in women with cancer of the uterine cervix without pelvic lymph node metastasis.

OBJECTIVES: To evaluate risk factors for recurrence of carcinoma of the uterine cervix among women who had undergone radical hysterectomy without pelvic lymph node metastasis, while taking into consideration not only the classical histopathological factors but also sociodemographic, clinical and treatment-related factors. STUDY DESIGN: This was an exploratory analysis on 233 women with carcinoma of the uterine cervix (stages IB and IIA) who were treated by means of radical hysterectomy and pelvic lymphadenectomy, with free surgical margins and without lymph node metastases on conventional histopathological examination. Women with histologically normal lymph nodes but with micrometastases in the immunohistochemical analysis (AE1/AE3) were excluded. Disease-free survival for sociodemographic, clinical and histopathological variables was calculated using the Kaplan-Meier method. The Cox proportional hazards model was used to identify the independent risk factors for recurrence. RESULTS: Twenty-seven recurrences were recorded (11.6%), of which 18 were pelvic, four were distant, four were pelvic+distant and one was of unknown location. The five-year disease-free survival rate among the study population was 88.4%. The independent risk factors for recurrence in the multivariate analysis were: postmenopausal status (HR 14.1; 95% CI: 3.7-53.6; P<0.001), absence of or slight inflammatory reaction (HR 7.9; 95% CI: 1.7-36.5; P=0.008) and invasion of the deepest third of the cervix (HR 6.1; 95% CI: 1.3-29.1; P=0.021). Postoperative radiotherapy was identified as a protective factor against recurrence (HR 0.02; 95% CI: 0.001-0.25; P=0.003). CONCLUSION: Postmenopausal status is a possible independent risk factor for recurrence even when adjusted for classical prognostic factors (such as tumour size, depth of tumour invasion, capillary embolisation) and treatment-related factors (period of treatment and postoperative radiotherapy status).