Clonal relationship and directionality of progression of synchronous endometrial and ovarian carcinomas in patients with DNA mismatch repair-deficiency associated syndromes.

Sporadic synchronous endometrial (ECs) and ovarian cancers (OCs), although clinically considered to be independent primaries, have been shown to be clonally related and likely constitute metastases from each other. We sought to define whether synchronous ECs/OCs in patients with DNA mismatch repair (MMR)-deficiency syndromes would be clonally related. We subjected synchronous ECs/OCs from four patients (LS3-LS6) with clinically confirmed Lynch syndrome (LS) and one patient with constitutional mismatch repair-deficiency syndrome (CMMRD) to massively parallel sequencing targeting 468 cancer-related genes. Somatic mutations, copy number alterations (CNAs), clonal relatedness and clonal decomposition analyses were performed using previously described bioinformatics methods. All synchronous ECs/OCs analyzed were considered independent primaries based on clinicopathologic criteria. Sequencing analysis revealed that the ECs/OCs of three cases (LS2-CMMRD, L3, L4) harbored similar repertoires of somatic mutations and CNAs and were clonally related. In these three cases, a subset of subclonal mutations in the EC became clonal in the OC, suggesting that the EC was likely the substrate from which the OC developed. LS5's EC/OC had distinct mutational profiles but shared specific CNAs. In contrast, LS6's EC/OC harbored distinct somatic mutations and lacked CNAs, consistent with each tumor constituting an independent primary lesion. In LS5 and LS6, PTEN mutations and PTEN loss of protein expression were found to be restricted to the EC. Finally, re-analysis of sequencing data of sporadic synchronous ECs/OCs supported the observations made in the current study that the directionality of progression is likely from the endometrium to the ovary. In conclusion, contrary to sporadic synchronous ECs/OCs, which are almost invariably clonally related, ECs/OCs simultaneously involving the uterus and ovary in LS patients may represent distinct primary tumors. A subset of MMR-deficiency syndrome-related synchronous ECs/OCs, however, may originate from a single primary tumor at variance with their clinical diagnosis, with the endometrium being the likeliest site of origin.

Mesonephric and mesonephric-like carcinomas of the female genital tract: molecular characterization including cases with mixed histology and matched metastases.

Mesonephric carcinoma of the cervix is a rare tumor derived from Wolffian remnants. Mesonephric-like carcinomas of the ovary and endometrium, while morphologically similar, do not have obvious Wolffian derivation. Here, we sought to characterize the repertoire of genetic alterations in primary mesonephric and mesonephric-like carcinomas, in the distinct histologic components of mixed cases, as well as in matched primary tumors and metastases. DNA from microdissected tumor and normal tissue from mesonephric carcinomas (cervix, n = 8) and mesonephric-like carcinomas (ovarian n = 15, endometrial n = 13) were subjected to sequencing targeting 468 cancer-related genes. The histologically distinct components of four cases with mixed histology and four primary tumors and their matched metastases were microdissected and analyzed separately. Mesonephric-like carcinomas were underpinned by somatic KRAS mutations (25/28, 89%) akin to mesonephric carcinomas (8/8, 100%), but also harbored genetic alterations more frequently reported in Müllerian tumors. Mesonephric-like carcinomas that lacked KRAS mutations harbored NRAS (n = 2, ovary) or BRAF (n = 1, endometrium) hotspot mutations. PIK3CA mutations were identified in both mesonephric-like (8/28, 28%) and mesonephric carcinomas (2/8, 25%). Only mesonephric-like tumors harbored CTNNB1 hotspot (4/28, 14%) and PTEN (3/13, 23%) mutations. Copy number analysis revealed frequent gains of chromosomes 1q and 10 in both mesonephric (87% 1q; 50% chromosome 10) and mesonephric-like tumors (89% 1q; 43% chromosome 10). Chromosome 12 gains were more frequent in ovarian mesonephric-like carcinomas, and losses of chromosome 9 were more frequent in mesonephric than in mesonephric-like carcinomas (both p = 0.01, Fisher's exact test). The histologically distinct components of four mixed cases were molecularly related and shared similar patterns of genetic alterations. The progression from primary to metastatic lesions involved the acquisition of additional mutations, and/or shifts from subclonal to clonal mutations. Our findings suggest that mesonephric-like carcinomas are derived from a Müllerian substrate with differentiation along Wolffian/mesonephric lines.

Histologic spectrum of polymorphous adenocarcinoma of the salivary gland harbor genetic alterations affecting PRKD genes.

Polymorphous adenocarcinoma (PAC) and cribriform adenocarcinoma of (minor) salivary gland (CASG) are salivary gland tumors with overlapping spectrum of morphology. Whether these represent distinct entities or a histologic spectrum of the same tumor remains contentious. PACs harbor recurrent PRKD1 E710D hotspot mutations in >70% of cases, whereas 80% of CASGs display rearrangements involving PRKD1, PRKD2, or PRKD3 (PRKD1/2/3). We studied the molecular and morphologic features of 37 PACs/CASGs, seeking to identify the associations among genotype, histologic phenotype, and classification. DNA was subjected to Sanger sequencing analysis of the PRKD1 hotspot locus. Fluorescence in situ hybridization (FISH) analysis for PRKD1/2/3 was performed using dual-color break-apart probes. Tumors were classified into four categories as described previously: PAC, CASG, tumor with indeterminate features (TIF), and tumor with a predominant papillary pattern (TPPP). PRKD1 E710D hotspot mutations were identified in 56%, 20%, 43% and 0% of PACs, CASGs, TIFs, and TPPPs, respectively. FISH demonstrated PRKD1/2/3 rearrangements in 13%, 78%, 36%, and 75% of PACs, CASGs, TIFs, and TPPPs, respectively. Histologically, fusion-positive tumors were associated with a high percentage of papillary growth, low percentage of single filing arrangement, a propensity of base of tongue location, and frequent (50%) lymph node metastasis, compared with the mutation-related tumors which had negligible nodal metastasis risk. Our results demonstrated that (1) PACs/CASGs are underpinned by genetic alterations affecting PRKD genes; (2) despite the associations between PAC and PRKD1 hotspot mutations and CASG and PRKD1/2/3 fusion, such distinction is not absolute; and (3) there is of a novel genotypic-phenotypic association whereby fusion-positive tumors are usually located in the base of the tongue, show papillary architecture and have a high risk of nodal metastasis. Genetic analysis of PRKD genes appears to be useful characterizing this spectrum of tumors, not only histologically but also clinically identifying those tumors with high risk of nodal metastasis.

Risk Factors for Persistence or Recurrence of High-Grade Cervical Squamous Intraepithelial Lesions.

OBJECTIVES: to evaluate whether the colposcopic lesion size , age, kind of surgery, the status of the surgical margins and the expression of the p16 and Ki-67 immunomarkers are risk factors for persistence or recurrence of the lesion. METHODS: a cross-sectional, observational, retrospective study of patients submitted to cold knife conization (CKC) or the loop electrosurgical excision procedure for cervical intraepithelial neoplasia 2 or 3. The colposcopic lesion size, age, surgical method, involvement of the surgical margins, and p16/Ki-67 immunomarker expression were analyzed in relation to lesion persistence and recurrence. RESULTS: seventy-one women were treated with cold knife conization and 200 were treated with loop electrosurgical excision. Of these, 95 had cervical intraepithelial neoplasia 2, 173 had cervical intraepithelial neoplasia 3, 183 had free surgical margins, 76 had compromised margins, and 12 showed damage by processing artifact or fragments. Among the 76 cases with positive margins, 55, 11, and 10 showed endocervical margin involvement, ectocervical margin involvement, and both endocervial and ectocervical margin involvement, respectively. Of the 264 followed-up patients, 38 had persistent or recurrent disease. A multiple logistic regression indicated that positive endocervical margins are the only independent risk factor for the persistence/recurrence of cervical intraepithelial neoplasia. No significant association was identified between the colposcopic lesion size, age, surgery type, or p16/Ki-67 immunomarker expression and lesion persistence or recurrence.

High-throughput proteomics of breast cancer subtypes: Biological characterization and multiple candidate biomarker panels to patients' stratification.

BACKGROUND AND AIMS: The actual classification of breast tumors in subtypes represents an attempt to stratify patients into clinically cohesive groups, nevertheless, clinicians still lack reproducible and reliable protein biomarkers for breast cancer subtype discrimination. In this study, we aimed to access the differentially expressed proteins between these tumors and its biological implications, contributing to the subtype's biological and clinical characterization, and with protein panels for subtype discrimination. METHODS: In our study, we applied high-throughput mass spectrometry, bioinformatic, and machine learning approaches to investigate the proteome of different breast cancer subtypes. RESULTS: We identified that each subtype depends on different protein expression patterns to sustain its malignancy, and also alterations in pathways and processes that can be associated with each subtype and its biological and clinical behaviors. Regarding subtype biomarkers, our panels achieved performances with at least 75% of sensibility and 92% of specificity. In the validation cohort, the panels obtained acceptable to outstanding performances (AUC = 0.740 to 1.00). CONCLUSIONS: In general, our results expand the accuracy of breast cancer subtypes' proteomic landscape and improve the understanding of its biological heterogeneity. In addition, we identified potential protein biomarkers for the stratification of breast cancer patients, improving the repertoire of reliable protein biomarkers. SIGNIFICANCE: Breast cancer is the most diagnosed cancer type worldwide and the most lethal cancer in women. As a heterogeneous disease, breast cancer tumors can be classified into four major subtypes, each presenting particular molecular alterations, clinical behaviors, and treatment responses. Thus, a pivotal step in patient management and clinical decisions is accurately classifying breast tumor subtypes. Currently, this classification is made by the immunohistochemical detection of four classical markers (estrogen receptor, progesterone receptor, HER2 receptor, and the Ki-67 index); however, it is known that these markers alone do not fully discriminate the breast tumor subtypes. Also, the poor understanding of the molecular alterations of each subtype leads to a challenging decision-making process regarding treatment choice and prognostic determination. This study, through high-throughput label-free mass-spectrometry data acquisition and downstream bioinformatic analysis, advances in the proteomic discrimination of breast tumors and achieves an in-depth characterization of the subtype's proteomes. Here, we indicate how the variations in the subtype's proteome can influence the tumor's biological and clinical differences, highlighting the variation in the expression pattern of oncoproteins and tumor suppressor proteins between subtypes. Also, through our machine-learning approach, we propose multi-protein panels with the potential to discriminate the breast cancer subtypes. Our panels achieved high classification performance in our cohort and in the independent validation cohort, demonstrating their potential to improve the current tumor discrimination system as complements to the classical immunohistochemical classification.

LncRNA-SNPs in a Brazilian Breast Cancer Cohort: A Case-Control Study.

Long noncoding RNAs (lncRNAs) are a class of non-coding RNAs that contain more than 200 nucleotides and exhibit a versatile regulatory capacity. Genomic alterations in lncRNAs have already been investigated in several complex diseases, including breast cancer (BC). BC is a highly heterogeneous disease and is the most prevalent cancer type among women worldwide. Single nucleotide polymorphisms (SNPs) in lncRNA regions appear to have an important role in BC susceptibility; however, little is known about lncRNA-SNPs in the Brazilian population. This study used Brazilian tumor samples to identify lncRNA-SNPs with a biological role in BC development. We applied a bioinformatic approach intersecting lncRNAs that are differentially expressed in BC tumor samples using The Cancer Genome Atlas (TCGA) cohort data and looked for lncRNAs with SNPs associated with BC in the Genome Wide Association Studies (GWAS) catalog. We highlight four lncRNA-SNPs-rs3803662, rs4415084, rs4784227, and rs7716600-which were genotyped in Brazilian BC samples in a case-control study. The SNPs rs4415084 and rs7716600 were associated with BC development at higher risk. These SNPs were also associated with progesterone status and lymph node status, respectively. The rs3803662/rs4784227 haplotype GT was associated with BC risk. These genomic alterations were also evaluated in light of the lncRNA's secondary structure and gain/loss of miRNA binding sites to better understand its biological functions. We emphasize that our bioinformatics approach could find lncRNA-SNPs with a potential biological role in BC development and that lncRNA-SNPs should be more deeply investigated in a highly heterogeneous disease population.

Colposcopic Findings and Diagnosis in Low-Income Brazilian Women with ASC-H pap Smear Results.

OBJECTIVE: To determine the accuracy of colposcopy findings in diagnosing cervical intraepithelial neoplasia (CIN) in women with an atypical squamous cells, cannot exclude high-grade squamous intraepithelial lesion (ASC-H) pap smear result and analyze whether the prevalence of HSIL and cancer correlates with sociodemographic risk factors and specific colposcopic findings. METHODS: Colposcopic findings and sociodemographic risk factors were analyzed as possible predictors of a CIN 2 or worse diagnosis in women with an ASC-H pap smear result. RESULTS: Accuracy of the colposcopic impression was 92%, sensitivity was 91.6%, and specificity was 93.1%, with a positive predictive value of 96.4% and negative predictive value of 84.3%. Diagnosis of CIN 2 or worse was more frequent in patients with a previous history of cervical dysplasia and pre-menopausal patients. Identification of major colposcopic findings, dense acetowhite epithelium, coarse mosaicism, and punctuation correlated significantly with CIN 2 or worse. CONCLUSION: Colposcopy performed by an experienced examiner can accurately differentiate patients with CIN 1 or less from patients with CIN 2 or worse. Diagnosis of CIN 2 or worse was more frequent in patients with a previous history of cervical dysplasia and pre-menopausal patients. The degree of acetowhite changes was the best colposcopic feature to predict CIN2 or worse.

OBJETIVO: Determinar a acurácia dos achados colposcópicos no diagnóstico das neoplasias intraepiteliais cervicais (NIC) em mulheres com resultado de exame citopatológico de células escamosas atípicas de significado indeterminado não podendo excluir lesão intraepitelial de alto grau (ASC-H) e analisar a correlação entre a prevalência de HSIL ou câncer com fatores de risco sociodemográficos e achados colposcópicos específicos. MéTODOS:: Os achados colposcópicos, e os fatores de risco sociodemográficos foram analisados como possíveis preditores de diagnóstico NIC 2 ou mais grave em mulheres com resultado de exame citopatológico ASC-H. RESULTADOS: A acurácia da impressão colposcópica foi de 92%, sensibilidade foi 91,6%, e a especificidade foi de 93,1%, com um valor preditivo de 96,4% e valor preditivo negativo de 84,3%. O diagnóstico de NIC 2 ou mais grave foi mais frequente em pacientes com história pregressa de displasia cervical e nas que não estavam na pós menopausa. A identificação de achados colposcópicos maiores, epitélio acetobranco denso, mosaico e pontilhados grosseiros se correlacionaram positivamente com o diagnóstico NIC 2 ou mais grave. CONCLUSãO:: A colposcopia realizada por um examinador experiente pode diferenciar com acurácia pacientes com NIC 1 ou menos grave de pacientes com NIC 2 ou mais grave. O diagnóstico de NIC 2 ou mais grave foi mais frequente em pacientes com história pregressa de displasia cervical e pacientes que estavam na pré menopausa. A densidade da acetorreação foi o melhor preditor colposcópico para NIC 2 ou mais grave.

Genomic Determinants of Early Recurrences in Low-Stage, Low-Grade Endometrioid Endometrial Carcinoma.

Low-stage, low-grade endometrioid endometrial carcinoma (EEC), the most common histologic type of endometrial cancer, typically has a favorable prognosis. A subset of these cancers, however, displays an aggressive clinical course with early recurrences, including distant relapses. All statistical tests were 2-sided. Using a combination of whole-exome and targeted capture sequencing of 65 FIGO stage IA and IB grade 1 EECs treated with surgery alone, we demonstrate that chromosome 1q gain (odds ratio [OR] = 8.09, 95% confidence interval [CI] = 1.59 to 54.6; P = .02), PIK3CA mutation (OR = 9.16, 95% CI = 1.95 to 61.8; P = .01), and DNA mismatch repair-deficient molecular subtype (OR = 7.92, 95% CI = 1.44 to 87.6; P = .02) are independent predictors of early recurrences within 3 years in this patient population. Chromosome 1q gain was validated in an independent dataset of stage I grade 1 EECs subjected to whole-exome sequencing. Our findings expand on the repertoire of genomic parameters that should be considered in the evaluation of patients with low-stage, low-grade EEC.

Stromal MED12 exon 2 mutations in complex fibroadenomas of the breast.

AIMS: Here we explore the presence of mediator complex subunit 12 (MED12) exon 2 and telomerase reverse transcriptase (TERT) promoter hotspot mutations in complex fibroadenomas (CFAs) of the breast. METHODS: The stromal components from 18 CFAs were subjected to Sanger sequencing of MED12 exon 2 and the TERT promoter hotspot loci. The epithelial and stromal components of two MED12 mutated CFAs were subjected to laser capture microdissection, and Sanger sequencing of MED12 exon 2, TERT promoter and PIK3CA exons 9 and 20, separately. RESULTS: MED12 exon 2 mutations were identified in the stroma of 17% of CFAs. The analyses of epithelial and stromal components, microdissected separately, revealed that MED12 mutations were restricted to the stroma. No TERT promoter or PIK3CA mutations in exons 9 and 20 were detected in analysed CFAs. CONCLUSIONS: Like conventional fibroadenomas, MED12 exon 2 mutations appear to be restricted to the stromal component of CFAs, supporting the notion that CFAs are stromal neoplasms.

Comprehensive analysis of the large and small ribosomal proteins in breast cancer: Insights on proteomic and transcriptomic expression patterns, regulation, mutational landscape, and prognostic significance.

Several evidence has demonstrated the involvement of the ribosomal proteins (RPs) in many malignancies, however, the function and clinical relevance of the RPs in breast cancer remains unclear. The present study aims to contribute to the understanding of the role of the RPs in breast tumorigenesis and its clinical implications in the field of biomarker discovery and outcome prediction. We investigated the proteomic and transcriptomic expression of the RPs in non-tumor and tumor tissues of different breast cancer subtypes, and integrated bioinformatics approaches and online databases to comprehensively evaluate the potential functions, regulatory networks, mutational landscape, and prognostic values of the ribosomal proteins in breast cancer. Our results show that 33 RPs have deregulated expression in breast cancer and its subtypes and that 26 RPs have potential as prognostic markers in a subtype-dependent way, with mutations in RP genes being frequent in breast tumors and related to overall survival and relapse-free status. Our RP gene regulatory network indicates the transcription factors MYC, ETS1, and SPI1, and the miRNAs has-let-7c-5p, has-mir-20b-5p, and has-mir-4668-3p as regulators of the RPs expression in breast cancer. The RPs were associated with several clinicopathological parameters of breast cancer and predicted to be involved in ribosomal-independent mechanisms such as regulation of the SLITS-ROBO pathway. This study comprehensively investigated the ribosomal proteins in breast cancer, suggesting that the RPs have clinical potential as biomarkers of diagnostic and prognostic, also providing an in-depth view of the RPs significance in breast cancer.

Prognostic significance of CD133 and ABCB5 expression in papillary thyroid carcinoma.

Expression of CD133 and ABCB5 is associated with tumor aggressiveness, but evidence in papillary thyroid cancer (PTC) is lacking. We correlated CD133 and ABCB5 expression with pathological characteristics and factors of worse prognosis in PTC. Samples of 119 PTCs and 40 controls (goiters) were distributed in 8 tissue microarray blocks and evaluated with immunohistochemistry using anti-CD133 and anti-ABCB5 antibodies. The expression of each marker alone and combined was analyzed against pathological characteristics and factors of worse prognosis in PTC. Expression of CD133 alone (19 tumors, 16.0%) was more frequent in patients with versus without lymph node metastases (P=0.024). Expression of ABCB5 alone (n=95, 83.3%) was associated with larger tumor size (P=0.045). CD133-ABCB5 coexpression was not associated with pathological characteristics or factors of worse prognosis in PTC.

Does human papillomavirus play a role in endometrial carcinogenesis?

SUMMARY: To determine the relationship between human papillomavirus (HPV) and endometrial carcinogenesis by comparing data from women with endometrial carcinoma to those of women with normal endometrial tissue. The survey was conducted for 100 women (50 with endometrial carcinoma and 50 with normal endometrial tissue) through HPV-DNA testing of paraffin-embedded endometrial tissue sections by polymerase chain reaction. Age, cigarette consumption, squamous differentiation and tumor grade, endometrium trophism, and HPV types detected in endometrial tissues were studied. HPV estimated odds ratio was similar in endometrial carcinoma and in normal endometrial tissue. The presence of HPV was not associated with age, tobacco abuse, endometrial histology status, squamous differentiation, or tumoral grade. DNA sequences of HPV types 16 and 18 were the most frequently detected in both groups. An association between HPV and endometrial carcinoma was not observed.

HER2 assessment by bright-field dual in situ hybridization in cell blocks of recurrent and metastatic breast carcinoma.

BACKGROUND: Breast cancer recurrences or metastases often are diagnosed using cytology material. Cell blocks (CBs) with adequate cellularity are crucial for the determination of accurate hormonal and human epidermal growth factor receptor 2 (HER2) status and to guide treatment. In the current study, the authors evaluated the concordance of HER2 status between bright-field dual in situ hybridization (DISH), fluorescence in situ hybridization (FISH), and HER2 immunohistochemistry (IHC) performed on formalin-fixed CBs of recurrent and metastatic breast cancers. METHODS: The authors searched for patients who had breast carcinoma recurrences or metastases diagnosed between 2010 and 2018 by fine-needle aspiration or by the drainage of body cavity fluids with HER2 IHC and/or FISH performed on formalin-fixed CBs. Cases with adequate tumor cellularity (>50 cells) were selected. HER2 DISH was performed on all CBs. HER2 status of the primary breast carcinoma was recorded. RESULTS: Formalin-fixed CBs were identified from 30 patients with breast cancer recurrences and metastases in axillary lymph nodes (LNs) (5 patients), mediastinal LNs (8 patients), internal mammary LNs (1 patient), supraclavicular LNs (2 patients), portocaval LNs (1 patient), chest wall (3 patients), pleural fluid (3 patients), bone (4 patients), liver (2 patients), and lung (1 patient). All cases had HER2 IHC performed at the study institution and were scored by breast pathologists according to the American Society of Clinical Oncology/College of American Pathologists guidelines. The HER2 DISH results demonstrated 100% concordance (30 of 30 cases) with the concurrent IHC and/or FISH. CONCLUSIONS: All methods of HER2 evaluation were found to accurately identify the amplification status. DISH can be used in tandem with IHC as a reflex assay instead of FISH and is an efficient and reliable method with which to determine HER2 amplification in formalin-fixed CBs.

Endometrial Cancers in BRCA1 or BRCA2 Germline Mutation Carriers: Assessment of Homologous Recombination DNA Repair Defects.

PURPOSE: Endometrial cancer (EC) is not considered a component of the hereditary breast and ovarian cancer syndrome but can arise in patients with germline BRCA1/2 (gBRCA1/2) mutations. Biallelic BRCA1/2 alterations are associated with genomic features of homologous recombination DNA repair deficiency (HRD) in cancer. We sought to determine if ECs in gBRCA1/2 mutation carriers harbor biallelic alterations and/or features of HRD. METHODS: Of 769 patients with EC who underwent germline panel testing, 10 pathogenic gBRCA1/2 mutation carriers were identified, and their tumor- and normal-derived DNA was subjected to massively parallel sequencing targeting at least 410 cancer-related genes. Three gBRCA1/2-associated ECs were identified in 232 ECs subjected to whole-exome sequencing by The Cancer Genome Atlas. Somatic mutations, copy number alterations, loss of heterozygosity, microsatellite instability (MSI), and genomic HRD features were assessed. RESULTS: Of the 13 patients included who had EC, eight harbored pathogenic gBRCA1 mutations and five harbored gBRCA2 mutations. Eight (100%) and two (40%) ECs harbored biallelic BRCA1 and BRCA2 alterations through loss of heterozygosity of the wild-type allele. All ECs harbored somatic TP53 mutations. One monoallelic/sporadic gBRCA2-associated EC had MLH1 promoter methylation and was MSI high. High large-scale state transition scores, a genomic feature of HRD, were found only in ECs with bi- but not monoallelic BRCA1/2 alterations. The Signature Multivariate Analysis HRD signature Sig3 was enriched in biallelic gBRCA1/2 ECs, and the three ECs from The Cancer Genome Atlas with BRCA1 biallelic alterations subjected to whole-exome sequencing displayed a dominant HRD-related mutational signature 3. CONCLUSION: A subset of gBRCA1/2-associated ECs harbor biallelic BRCA1/2 alterations and genomic features of HRD, which may benefit from homologous recombination-directed treatment regimens. ECs in BRCA2 mutation carriers might be sporadic and even MSI high, and may potentially benefit from immune-checkpoint inhibition.

Recurrent MED12 exon 2 mutations in benign breast fibroepithelial lesions in adolescents and young adults.

AIMS: Most benign breast fibroepithelial lesions (FEL) in adults harbour recurrent somatic MED12 exon 2 mutations and rare TERT promoter hotspot mutations. We sought to determine the frequency of MED12 exon 2 and TERT promoter hotspot mutations in fibroadenomas (FA) and benign phyllodes tumours (BePT) in adolescents and young adults. METHODS: DNA from 21 consecutive FAs and eight consecutive BePTs in adolescents and young adults was subjected to Sanger sequencing of the exon 2 of MED12 and the TERT promoter hotspot locus. RESULTS: We identified MED12 exon 2 mutations in 62% and 88% of FAs and BePTs, respectively, and no TERT promoter hotspot mutations. The majority of the MED12 exon 2 mutations identified were in-frame deletions (60%). CONCLUSIONS: As in adults, benign FELs in juvenile patients harbour recurrent MED12 exon 2 mutations.

Risk Factors for Persistence or Recurrence of High-Grade Cervical Squamous Intraepithelial Lesions / Fatores de risco na persistência ou recidiva da Lesão escamosa intraepitelial de alto grau (LIEAG)

ABSTRACT Objectives: to evaluate whether the colposcopic lesion size , age, kind of surgery, the status of the surgical margins and the expression of the p16 and Ki-67 immunomarkers are risk factors for persistence or recurrence of the lesion. Methods: a cross-sectional, observational, retrospective study of patients submitted to cold knife conization (CKC) or the loop electrosurgical excision procedure for cervical intraepithelial neoplasia 2 or 3. The colposcopic lesion size, age, surgical method, involvement of the surgical margins, and p16/Ki-67 immunomarker expression were analyzed in relation to lesion persistence and recurrence. Results: seventy-one women were treated with cold knife conization and 200 were treated with loop electrosurgical excision. Of these, 95 had cervical intraepithelial neoplasia 2, 173 had cervical intraepithelial neoplasia 3, 183 had free surgical margins, 76 had compromised margins, and 12 showed damage by processing artifact or fragments. Among the 76 cases with positive margins, 55, 11, and 10 showed endocervical margin involvement, ectocervical margin involvement, and both endocervial and ectocervical margin involvement, respectively. Of the 264 followed-up patients, 38 had persistent or recurrent disease. A multiple logistic regression indicated that positive endocervical margins are the only independent risk factor for the persistence/recurrence of cervical intraepithelial neoplasia. No significant association was identified between the colposcopic lesion size, age, surgery type, or p16/Ki-67 immunomarker expression and lesion persistence or recurrence.

RESUMO Objetivos: avaliar se o status das margens, idade, tamanho da lesão colposcópica, tipo de cirurgia e expressão dos marcadores p16/Ki-67 são fatores de risco na persistência ou recidiva da LIEAG. Métodos: um estudo de corte transversal, observacional com coleta de dados retrospectivos de pacientes submetidas a conização a frio (CF) ou exérese da zona de transformação por cirurgia de alta frequência EZT por NIC2/3. Foram analisados os seguintes fatores em relação a persistência ou recidiva: comprometimento das margens, idade, tamanho da lesão, tipo de cirurgia e coexpressão dos imunomarcadores p16 e Ki-67. Resultados: 271 mulheres tratadas com CF (71) e EZT (200), onde 95 apresentavam NIC 2 e 173 NIC 3, 183 apresentaram margens cirúrgicas livres, 76 comprometidas e 12 prejudicadas por artefatos ou fragmentação. Das 76 pacientes com margens comprometidas, 55 foram endocervical, 11 ectocervical e 10 ambas as margens. Das 264 pacientes que tiveram seguimento, 38 persistiram ou recidivaram a doença. A regressão logística múltipla indicou ser a margem endocervical comprometida o único fator independente de risco de persistência/recorrência da NIC (p<0,001). Não houve associação significativa entre a idade, o tamanho da lesão colposcópica, o tipo de cirurgia e a expressão dos imunomarcadores p16/Ki-67 e a persistência ou recorrência da doença. Conclusão: entre os fatores estudados associados com persistência ou recorrência, somente a margem endocervical comprometida provou ser significativamente um fator risco para persistência ou recorrência da lesão.

Cancer stem cell markers ALDH1 and CD44+/CD24- phenotype and their prognosis impact in invasive ductal carcinoma.

Breast cancer is a very heterogeneous disease. The intrinsic molecular subtypes can explain the intertumoral heterogeneity and the cancer stem cell (CSC) hypothesis can explain the intratumoral heterogeneity of this kind of tumor. CD44+/CD24- phenotype and ALDH1 expression are the major CSC markers described in invasive breast cancer. In the present study, 144 samples of invasive breast carcinoma, no special type were distributed in 15 tissue microarrays (TMA) and then evaluated for expression of the CD44+/CD24- phenotype and ALDH1 to understand the importance of these CSC markers and the clinical aspects of breast cancer. The samples were classified into four molecular subtypes according to clinicopathological criteria: Luminal A, Luminal B, HER2, and Basal-like. A statistical association was found between the molecular subtypes and the CSC markers, with HER2 the most frequent subtype for both markers. ALDH1 was also associated with other poor prognostic variables, such as a high histological grade and larger tumors, but it was not associated with the patients' prognosis in this sample and nor was the CD44+/CD24- phenotype in a multivariate analysis. There are still many controversies about the role of these markers in breast cancer molecular subtypes. The identification of these populations of cells, through immunohistochemical markers, can help to better understand the CSC theory in clinical practice and, in the near future, contribute to developing new target therapies.

Concordância Interobservador no Diagnóstico Citológico de Atipia Escamosa de Significado Indeterminado Favorecendo Lesão de Alto Grau e de Lesão / Intraepitelial Escamosa de Alto Grau nas Lesões do Colo Uterino Interobserver Agreement in Cytological Diagnosis of Atypical Squamous Cell, Cannot Exclude a High-Grade Lesion and High-Grade Squamous Intraepithelial Lesion in Cervical Lesions / Concordancia Interobservador en el Diagnóstico Citológico de Atipia Escamosa de Significado Indeterminado Favoreciendo Lesión de Alto Grado y Lesión Intraepitelial Escamosa de Alto Grado en Lesiones del Cuello Uterino

Introdução: O exame de Papanicolau é uma importante ferramenta na triagem do carcinoma do colo uterino. O diagnóstico citológico de atipias celulares escamosas de significado indeterminado favorecendo lesão de alto grau (ASC-H) é a categoria de menor concordância interobservador. Objetivo: Avaliar o grau de concordância interobservador para os diagnósticos de ASC-H e de lesões intraepiteliais escamosas de alto grau (LIEAG) em um hospital terciário e avaliar a capacidade do diagnóstico de ASC-H para predizer lesões de maior grau. Método: Foram coletadas lâminas de pacientes atendidas entre 2007 e 2015 no Serviço de Anatomia Patológica do hospital, com diagnósticos originais de ASC-H ou LIEAG realizados pelo mesmo patologista, colposcopia e biópsia, quando indicadas, pelo mesmo ginecologista. Essas citologias foram posteriormente revisadas por outros dois patologistas separadamente e às cegas. Ambos tiveram acesso a dados sobre idade no momento do diagnóstico para reproduzir o diagnóstico da prática clínica. Resultados: Houve 65,1% de lâminas listadas com ASC-H e 34,9% com LIEAG. As duas revisões concordaram concomitantemente com o diagnóstico original em 54,7%. Os índices kappa para os dois diagnósticos e somente para ASC-H foram, respectivamente, 0,46 e 0,49 (concordâncias moderadas). Das lâminas originalmente interpretadas como ASC-H, 68,3% resultaram em lesões de maior grau na histologia. Conclusão: Os dados mostraram uma concordância moderada entre os patologistas para o diagnóstico de ASC-H. É importante destacar que o diagnóstico de ASC-H correspondeu à lesão de maior grau de malignidade na histologia, demonstrando que essas lesões devem ser seguidas clinicamente como LIEAG

Introduction: The Papanicolaou test is an important screening exam for cervical carcinoma. The cytological diagnosis of undetermined atypical squamous cells favoring high-grade lesion (ASC-H) is the category with the least interobserver concurrence. Objective: Evaluate the interobserver concurrence for the ASC-H and high-grade squamous intraepithelial lesions (HSIL) categories at a teaching hospital and to estimate ASC-H's capacity to predict higher grade lesions. Method: Smears from patients admitted from 2007 to 2015 whose original diagnosis was made by one pathologist, in addition to colposcopy and biopsy, when indicated, made by one gynecologist were collected in the Pathologic Anatomy Service of the hospital. The cytology was reviewed by two other pathologists separately and blindly. Both reviewers had access to data about age at the moment of the diagnosis in order to reproduce the clinical diagnosis. Results: There were 65.1% smears considered as ASC-H and 34.9%, as HSIL. The reviews concurred simultaneously with the original diagnosis in 54.7% of the cases. The kappa indexes for both categories and only for ASC-H were, respectively, 0.46 and 0.49 (moderate concurrence). 68.3% of the smears primarily described as ASC-H resulted in higher grade lesions in histology. Conclusion: The data showed a moderate concurrence between the pathologists for the ASC-H's diagnosis. It is important to highlight that ASC-H matched with higher grade lesions at the histology, needing follow-up as HSIL

Introducción: La prueba de Papanicolaou es un importante examen de detección del carcinoma del cuello uterino. El diagnostico citológico de las células escamosas atípicas, no se descarta una lesión de grado alto (ASC-H) es la categoría de menor acuerdo interobservador. Objetivo: Los objetivos de este estudio fueron evaluar el grado de concordancia interobservador para los diagnósticos de atipias escamosas de significado indeterminado favoreciendo lesión de alto grado (ASC-H) y de lesiones intraepiteliales escamosas de alto grado (LIEAG) en un hospital terciario de Curitiba (PR) y evaluar la capacidad del diagnóstico de ASC-H de predecir las lesiones de mayor grado. Método: Se recogieron del Servicio de Anatomía Patológica del hospital las láminas de pacientes atendidas entre 2007 y 2015, con diagnósticos originales de ASC-H o LIEAG realizados por el mismo patólogo y colposcopia y biopsia, cuando indicadas, por el mismo ginecólogo. Esas citologías fueron revisadas después por otros dos patólogos separadamente y a ciegas. Ambos tuvieron acceso a datos sobre edad en el momento del diagnóstico para reproducir el diagnóstico de la práctica clínica. Resultados: Hubo el 65,1% de las láminas señaladas con ASC-H y el 34,9%, con LIEAG. Las revisiones concordaron concomitantemente con el diagnóstico original en el 54,7%. Los índices kappa para los dos diagnósticos y solamente para ASC-H fueron, respectivamente, 0,46 y 0,49 (concordancias moderadas). De las láminas originalmente interpretadas como ASC-H, 68,3% resultaron en lesiones de mayor grado en la histología. Conclusión: Hubo una concordancia moderada entre los patólogos para la categoría ASC-H. Se destaca también la correspondencia de ASC-H con lesiones de mayor grado en la histología, lo que dirige su seguimiento clínico como LIEAG

[Extract of Passiflora edulis in the healing of colonic anastomosis in rats: a tensiometric and morphologic study]. / Extrato de Passiflora edulis na cicatrização de anastomose colônica em ratos: estudo morfológico e tensiométrico.

INTRODUCTION: Investigation of new substances with therapeutic effects have been done trying to isolate, extract or purify new compounds of vegetable origin. The Passiflora edulis (maracujá) species from the Plassifloracia family, originated from the tropical and subtropical regions of the american continent, is found all over Brazil. It is commonly used as a sedative, painkiller and anti-inflammatory drug and also for the treatment of skin wounds, lesions and Erisipelae. PURPOSE: To evaluate the wound healing in colonic anastomosis in rats that received an hydro-alcoholic extract of Passiflora edulis peri-operatively. METHOD: 40 wistar rats were used distributed into two groups of 20 rats each, named: Passiflora edulis group (GP) and control group (GC). The rats of each group were separated into two subgroups of 10 animals each and were evaluated on the 3rd and 7th postoperative days. The surgical procedure consisted of a section of the left colon, 5 cm above the peritoneal reflexion with preservation of the vascular elements. Intestinal continuity was restored by an end-to-end single layer anastomosis. The Passiflora edulis group received an intraperitoneal application of the hydro-alcoholic extract in the dosage of 250 mg/kg. The control-group received one intraperitoneal dose of a saline with the same volume of the GP. The parameters evaluated were: macroscopic aspects of the wall and abdominal cavity, perianastomotic (adherences), bursting pressure, inflammatory tissue reaction on the anastomotic wound. RESULTS: The macroscopic aspects did not differ between the groups. No rupture in the anastomotic wound was seen in any rat. Regarding the bursting pressure, it was noticed that the average pressure was significantly higher in the subgroup that received the Passiflora edulis extract on the 3rd day (P3) (42.6 +/- 17.8 mmHg vs. 25.4 +/- 14.1 mmHg, p=0.028), as compared to the control sub-group (C3). However, on the 7th day, bursting pressure was similar in both groups (p=0.447). Rats from the C7 sub-group had a mean bursting pressure of 203.0 +/- 50.0 mmHg vs. 187.3 +/- 39.5 mmHg in the C7 sub-group. In the histologic analysis the polymorphic nuclear cells were more frequent in the C3 group, with significant differences (p=0.034). The monomorphic nuclear cells (MMN) and the fibroblastic proliferation were more frequent in the P3 sub-group with a significant difference, p=0.02 to MMN, and p=0.001 to the fibroblastic proliferation. On the 7th day there was a significant difference in all histologic criteria stained by hematoxin-eosin and Masson Trichomic (p<0.05) in the sub-group that received the Passiflora edulis extracts. CONCLUSION: The peri-operative administration of the hydro-alcoholic extract of Passiflora edulis has a positive influence on the healing of colonic anastomosis in rats.

The role of Phosphatidylinositol 3 kinase (PI3K) and Cycloxygenase-2 (COX2) in carcinogenesis of colorectal polyps / O papel de PI3K e COX2 na carcinogênese de pólipos colorretais

ABSTRACT Objectives: Determine immunohistochemical expression of Phosphatase and tensin homolog (PTEN), Phosphatidylinositol 3 kinase (PI3K), Cycloxygenase-2 (COX2) and one proliferation marker (Ki67) in colorectal polyps and correlate with clinical and pathological data in search of carcinogenic pathways. Methods: The reports of 297 polyps diagnosed through endoscopy were reviewed for parameters including age, gender, prior colorectal cancer, the presence of multiple polyps, and polyps' location, appearance and size. Was conducted a microscopic morphometric computerized analysis of immunohistochemical expression using, the selected antibodies and correlated with clinical and pathological variables. Results: The tissue immunohistochemical expression was higher in right colon polyps for the proliferation marker and Phosphatidylinositol 3 kinase (p ≤ 0.0001 and 0.057 respectively). Cycloxygenase-2 and Phosphatase and tensin homolog demonstrated higher tissue immunoexpression in pedunculated polyps (p = 0.009 and 0.002 respectively). Cycloxygenase-2 exhibited higher immunoexpression in larger polyps (p = 0.005). Phosphatidylinositol 3 kinase, Cycloxygenase-2, Phosphatase and tensin homolog and the proliferation marker exhibited higher immunoexpression in high-grade dysplastic polyps (p = 0.031, 0.013, 0.044 and <0.001 respectively). Phosphatase and tensin homolog labeling was higher in polyps with high-grade dysplasia and lower in some of serrated lesions (p = 0.044). Conclusions: The greater expression of the proliferation marker and Phosphatidylinositol 3 kinase in the right colon may be related to right-sided colorectal carcinogenesis. The proliferation marker, Cycloxygenase-2 and Phosphatidylinositol 3 kinase results can be associated with progression of polyps to colorectal cancer. The higher Phosphatase and tensin homolog expression suggests its attempt to control the cell cycle.

RESUMO Objetivos: Determinar a expressão imuno-histoquímica de Fosfatase homóloga a tensina (PTEN), Fosfatidilinositol-3-cinase (PI3K), Ciclooxigenase-2 (COX2) e um marcador de proliferação (Ki67) em pólipos colorretais e correlacionar com dados clínicos e patológicos buscando sua correspondência na carcinogênese. Métodos: Revisados 297 pólipos diagnosticados através de endoscopia quanto a idade, gênero, história de câncer colorretal, número, localização, aparência e tamanho dos pólipos. Realizadas as avaliações morfométricas computadorizadas das expressões imuno-histoquímicas dos marcadores selecionados, que foram correlacionadas com variáveis clínicas e patológicas. Resultados: A expressão do marcador de proliferação e da Fosfatidilinositol-3-cinase foi maior nos pólipos do cólon direito (p = <0,0001 e 0.057 respectivamente). Ciclooxigenase-2 e Fosfatase homóloga a tensina demonstraram maior imunoexpressão em pólipos pediculados (p = 0,009 e 0,002, respectivamente). Ciclooxigenase-2 expressou mais em pólipos maiores (p = 0,005). Fosfatidilinositol-3-cinase, Ciclooxigenase-2, Fosfatase homóloga a tensina e o marcador de proliferação expressaram mais em pólipos com displasia de alto grau (p = 0,031, 0,013, 0,044 e <0,001, respectivamente). Fosfatase homóloga a tensina marcou mais pólipos com displasia de alto grau que lesões serrilhadas (p = 0,044). Conclusões: A maior expressão do marcador de proliferação e Fosfatidilinositol-3-cinase à direita pode estar relacionada à carcinogênese do lado direito do cólon. Os resultados do marcador de proliferação, Ciclooxigenase-2 e Fosfatidilinositol-3-cinase podem ser associados à progressão dos pólipos para câncer. A expressão aumentada de Fosfatase homóloga a tensina sugere tentativa de controle do ciclo celular.