Unveiling the RKIP and EGFR Inverse Relationship in Solid Tumors: A Case Study in Cervical Cancer.

Raf Kinase Inhibitor Protein (RKIP) is recognized as a bona fide tumor suppressor gene, and its diminished expression or loss is associated with the progression and poor prognosis of various solid tumors. It exerts multifaceted roles in carcinogenesis by modulating diverse intracellular signaling pathways, including those governed by HER receptors such as MAPK. Given the significance of HER receptor overexpression in numerous tumor types, we investigated the potential oncogenic relationship between RKIP and HER receptors in solid tumors. Through a comprehensive in silico analysis of 30 TCGA PanCancer Atlas studies encompassing solid tumors (10,719 samples), we uncovered compelling evidence of an inverse correlation between RKIP and EGFR expression in solid tumors observed in 25 out of 30 studies. Conversely, a predominantly positive association was noted for the other HER receptors (ERBB2, ERBB3, and ERBB4). In particular, cervical cancer (CC) emerged as a tumor type exhibiting a robust inverse association between RKIP and EGFR expression, a finding that was further validated in a cohort of 202 patient samples. Subsequent in vitro experiments involving pharmacological and genetic modulation of EGFR and RKIP showed that RKIP depletion led to significant upregulation of EGFR mRNA levels and induction of EGFR phosphorylation. Conversely, EGFR overactivation decreased RKIP expression in CC cell lines. Additionally, we identified a common molecular signature among patients depicting low RKIP and high EGFR expression and demonstrated the prognostic value of this inverse correlation in CC patients. In conclusion, our findings reveal an inverse association between RKIP and EGFR expression across various solid tumors, shedding new light on the underlying molecular mechanisms contributing to the aggressive phenotype associated with RKIP and EGFR in cervical cancer.

Molecular analysis of translocations associated with Ewing sarcoma: case report / Análise molecular de translocações associadas ao sarcoma de Ewing: relato de caso

ABSTRACT We report the case of a 25-year-old female patient having a large lesion in the right breast. The core biopsy revealed a high-grade fusocellular neoplasm. Immunostains show no evidence of a specific line of differentiation. The patient underwent radical surgery and adjuvant treatment. Due to the immunohistochemical profile and the rosette arrangement in fusocellular injury, a molecular evaluation of translocations associated to the Ewing sarcoma (ES) was made. However, translocations commonly associated with this disease [t(11;22)(q24;q12)] were not observed. The present report can contribute to the diagnostic investigation of similar cases and to the pre-test orientation of the molecular evaluation.

RESUMEN Presentamos el caso de una paciente de 25 años con lesión voluminosa en mama derecha. La biopsia de la lesión mostró neoplasia fusocelular de alto grado; la inmunohistoquímica no ha determinado la histogénesis de la lesión. La paciente fue sometida a la cirugía radical y al tratamiento adyuvante. Ante el perfil inmunohistoquímico y la formación en roseta de la lesión fusocelular, se ha realizado una evaluación molecular de translocaciones asociadas al sarcoma de Ewing (SE); sin embargo no han sido observadas translocaciones comúnmente asociadas a esa patología [t(11;22)(q24;q12)]. El presente reporte puede ayudar en la investigación diagnóstica en casos similares así como orientar la evaluación molecular.

RESUMO Relatamos o caso de uma paciente, 25 anos de idade, portadora de lesão volumosa em mama direita. A biópsia da lesão evidenciou neoplasia fusocelular de alto grau; a imuno-histoquímica não revelou a histogênese da lesão. A paciente foi submetida à cirurgia radical e ao tratamento adjuvante. Diante do perfil imuno-histoquímico e do arranjo em roseta em lesão fusocelular, foi realizada avaliação molecular de translocações associadas ao sarcoma de Ewing (SE); entretanto não foram observadas translocações comumente associadas a essa patologia [t(11;22)(q24;q12)]. O presente relato pode contribuir para a investigação diagnóstica de casos semelhantes, bem como para a orientação pré-teste da avaliação molecular.

HER Family Receptors are Important Theranostic Biomarkers for Cervical Cancer: Blocking Glucose Metabolism Enhances the Therapeutic Effect of HER Inhibitors.

Persistent HPV infection alone is not sufficient for cervical cancer development, which requires additional molecular alterations for tumor progression and metastasis ultimately leading to a lethal disease. In this study, we performed a comprehensive analysis of HER family receptor alterations in cervical adenocarcinoma. We detected overexpression of HER protein, mainly HER2, which was an independent prognostic marker for these patients. By using in vitro and in vivo approaches, we provided evidence that HER inhibitors, allitinib and lapatinib, were effective in reducing cervical cancer aggressiveness. Furthermore, combination of these drugs with glucose uptake blockers could overcome the putative HIF1-α-mediated resistance to HER-targeted therapies. Thus, we propose that the use of HER inhibitors in association with glycolysis blockers can be a potentially effective treatment option for HER-positive cervical cancer patients.

Reprogramming energy metabolism and inducing angiogenesis: co-expression of monocarboxylate transporters with VEGF family members in cervical adenocarcinomas.

BACKGROUND: Deregulation of cellular energetic metabolism was recently pointed out as a hallmark of cancer cells. This deregulation involves a metabolic reprogramming that leads to a high production of lactate. Lactate efflux, besides contributing for the glycolytic flux, also acts in the extracellular matrix, contributing for cancer malignancy, by, among other effects, induction of angiogenesis. However, studies on the interplay between cancer metabolism and angiogenesis are scarce. Therefore, the aim of the present study was to evaluate the metabolic and vascular molecular profiles of cervical adenocarcinomas, their co-expression, and their relation to the clinical and pathological behavior. METHODS: The immunohistochemical expression of metabolism-related proteins (MCT1, MCT4, CD147, GLUT1 and CAIX) as well as VEGF family members (VEGF-A, VEGF-C, VEGF-D, VEGFR-1, VEGFR-2 and VEGFR-3) was assessed in a series of 232 cervical adenocarcinomas. The co-expression among proteins was assessed and the expression profiles were associated with patients' clinicopathological parameters. RESULTS: Among the metabolism-related proteins, MCT4 and CAIX were the most frequently expressed in cervical adenocarcinomas while CD147 was the less frequently expressed protein. Overall, VEGF family members showed a strong and extended expression with VEGF-C and VEGFR-2 as the most frequently expressed and VEGFR-1 as the less expressed member. Co-expression of MCT isoforms with VEGF family members was demonstrated. Finally, MCT4 was associated with parametrial invasion and HPV18 infection, CD147 and GLUT1 with distant metastasis, CAIX with tumor size and HPV18 infection, and VEGFR-1 with local and lymphnode metastasis. CONCLUSIONS: The results herein presented provide additional evidence for a crosstalk between deregulating cellular energetics and inducing angiogenesis. Also, the metabolic remodeling and angiogenic switch are relevant to cancer progression and aggressiveness in adenocarcinomas.

RKIP inhibition in cervical cancer is associated with higher tumor aggressive behavior and resistance to cisplatin therapy.

Cervical cancer is one of the most common cancers in women worldwide, being high-risk group the HPV infected, the leading etiological factor. The raf kinase inhibitory protein (RKIP) has been associated with tumor progression and metastasis in several human neoplasms, however its role on cervical cancer is unclear. In the present study, 259 uterine cervix tissues, including cervicitis, cervical intraepithelial lesions and carcinomas, were analyzed for RKIP expression by immunohistochemistry. We found that RKIP expression was significantly decreased during malignant progression, being highly expressed in non-neoplastic tissues (54% of the samples; 73/135), and expressed at low levels in the cervix invasive carcinomas (∼15% (19/124). Following in vitro downregulation of RKIP, we observed a viability and proliferative advantage of RKIP-inhibited cells over time, which was associated with an altered cell cycle distribution and higher colony number in a colony formation assay. An in vitro wound healing assay showed that RKIP abrogation is associated with increased migratory capability. RKIP downregulation was also associated with an increased vascularization of the tumors in vivo using a CAM assay. Furthermore, RKIP inhibition induced cervical cancer cells apoptotic resistance to cisplatin treatment. In conclusion, we described that RKIP protein is significantly depleted during the malignant progression of cervical tumors. Despite the lack of association with patient clinical outcome, we demonstrate, in vitro and in vivo, that loss of RKIP expression can be one of the factors that are behind the aggressiveness, malignant progression and chemotherapy resistance of cervical cancer.

Molecular characterization of EGFR, PDGFRA and VEGFR2 in cervical adenosquamous carcinoma.

BACKGROUND: Adenosquamous carcinoma of the uterine cervix is an infrequent but aggressive subtype of cervical cancer. A better understanding of its biological behaviour is warranted to define more accurate prognosis and therapeutic targets. Currently, the blockage of receptor tyrosine kinase (RTKs) activity is an efficient therapeutic strategy for many different cancers. The objective of this study was to investigate EGFR, PDGFRA and VEGFR2 RTKs overexpression and activating gene mutations in a cohort of 30 adenosquamous carcinomas of the uterine cervix. METHODS: EGFR, PDGFRA and VEGFR2 immunohistochemistry was performed in all samples, followed by DNA isolation from the gross macroscopically dissection of the neoplastic area. Screening for EGFR (exons 18-21) and PDGFRA (exons 12, 14 and 18) mutations was done by PCR--single-strand conformational polymorphism (PCR-SSCP). RESULTS: Despite the presence of EGFR immunohistochemical positive reactions in 43% (13/30) of the samples, no EGFR activating mutations in the hotspot region (exons 18-21) were identified. A silent base substitution (CAG>CAA) in EGFR exon 20 at codon 787 (Q787Q) was found in 17 cases (56%). All PDGFRA immunohistochemical reactions were positive and consistently observed in the stromal component, staining fibroblasts and endothelial cells, as well as in the cytoplasm of malignant cells. No activating PDGFRA mutations were found, yet, several silent mutations were observed, such as a base substitution in exon 12 (CCA>CCG) at codon 567 (P567P) in 9 cases and in exon 18 (GTC>GTT) at codon 824 (V824V) in 4 cases. We also observed the presence of base substitutions in intron 14 (IVS14+3G>A and IVS14+49G>A) in two different cases, and in intron 18 (IVS18-50insA) in 4 cases. VEGFR2 positivity was observed in 22 of 30 cases (73.3%), and was significantly associated with lack of metastasis (p=0.038). CONCLUSION: This is the most extensive analysis of EGFR, PDGFRA and VEGFR2 in cervical adenosquamous carcinomas. Despite the absence of EGFR and PDGFRA activating mutations, the presence of overexpression of these three important therapeutic targets in a subset of cases may be important in predicting the sensitivity of adenosquamous carcinoma to specific anti-RTKs drugs.

Monocarboxylate transporters 1 and 4 are associated with CD147 in cervical carcinoma.

Due to the highly glycolytic metabolism of solid tumours, there is an increased acid production, however, cells are able to maintain physiological pH through plasma membrane efflux of the accumulating protons. Acid efflux through MCTs (monocarboxylate transporters) constitutes one of the most important mechanisms involved in tumour intracellular pH maintenance. Still, the molecular mechanisms underlying the regulation of these proteins are not fully understood. We aimed to evaluate the association between CD147 (MCT1 and MCT4 chaperone) and MCT expression in cervical cancer lesions and the clinico-pathological significance of CD147 expression, alone and in combination with MCTs. The series included 83 biopsy samples of precursor lesions and surgical specimens of 126 invasive carcinomas. Analysis of CD147 expression was performed by immunohistochemistry. CD147 expression was higher in squamous and adenocarcinoma tissues than in the non-neoplastic counterparts and, importantly, both MCT1 and MCT4 were more frequently expressed in CD147 positive cases. Additionally, co-expression of CD147 with MCT1 was associated with lymph-node and/or distant metastases in adenocarcinomas. Our results show a close association between CD147 and MCT1 and MCT4 expressions in human cervical cancer and provided evidence for a prognostic value of CD147 and MCT1 co-expression.

Increasing expression of monocarboxylate transporters 1 and 4 along progression to invasive cervical carcinoma.

Solid tumor cells are known to be highly glycolytic and, to prevent apoptosis by cellular acidosis, cells increase proton efflux through pH regulators, such as monocarboxylate transporters (MCTs). However, the role of these membrane proteins in solid tumor development and survival is not fully understood. We aimed to evaluate the expression of the MCT isoforms 1, 2, and 4 in a large series of cervical lesions (neoplastic and non-neoplastic) and assess its clinical-pathologic significance. The series analyzed included 29 chronic cervicitis, 30 low-grade squamous intraepithelial lesions, 32 high-grade squamous intraepithelial lesions, 49 squamous cell carcinomas, 51 adenocarcinomas, and 30 adenosquamous carcinomas of the uterine cervix. Analysis of the expression of MCT isoforms 1, 2, and 4 was performed by immunohistochemistry with specific antibodies. Immunoreactions were evaluated both qualitatively and semiquantitatively. We found a significant increase in MCT expression from preinvasive to invasive squamous lesions and from normal glandular epithelium to adenocarcinomas. This is the first study evaluating the significance of MCT expression in lesions of the uterine cervix, including invasive carcinomas, and the results found herein led us to believe that these membrane proteins are involved in the progression to invasiveness in uterine cervix carcinoma.

KIT activation in uterine cervix adenosquamous carcinomas by KIT/SCF autocrine/paracrine stimulation loops.

OBJECTIVES: Uterine adenosquamous carcinoma (ASC) is an uncommon, yet, one of the most aggressive cervical cancer subtype. The successful treatment of some tumors, such as gastrointestinal stromal tumors (GISTs), by anti-KIT inhibitors fosters the study of this receptor tyrosine kinase in other malignancies. In the present study, we intended to molecularly characterize KIT in ASC. METHODS: In a series of 30 cases, we studied KIT (CD117), KIT phosphorylated/activated form, as well as KIT ligand, stem cell factor (SCF), by immunohistochemistry. We further screened for KIT hotspot mutations (exon 9, 11, 13 and 17) by PCR-SSCP and for KIT gene amplification by Quantitative real-time PCR in CD117 positive cases. RESULTS: We observed CD117 expression in approximately 13% of cases, with approximately 7% co-expressing SCF, which resulted in KIT phosphorylation/activation. No KIT activating mutations or gene amplification were found, despite the presence of 4q aneuploidy in one case. CONCLUSIONS: This is the first study assessing KIT activation and molecular alterations in a large series of rare ASC. Our findings showed the absence of KIT molecular alterations and suggested the presence of KIT activation in a small proportion of cases through KIT/SCF co-expression.

How accurate is cytological diagnosis of cervical glandular lesions?

Gynecological cytology has some inaccurate morphological categorization and poor interobserver reproducibility especially for glandular lesions. Liquid-based cytology (LBC) preparations are presumed to reduce artifacts that interfere in diagnosis performance, but its value to correctly identify glandular alterations has not been sufficiently reported. The objective of this study was to compare the diagnostic performance and interobserver agreement of LBC and conventional Pap smear to identify histologically confirmed glandular lesions according to five cytologists. Sensitivity ranged from 55.8 to 73.1% and 32.7 to 48.1% for Pap smear and LBC, respectively. Specificity ranged from 66.1 to 87.1% and 69.4 to 94.4%, respectively. In general, agreement between pairs of cytologists was poor with kappa-values around 0.45. In conclusion, relying entirely on cervical cytology to rule out glandular lesions may be risky. The use of HPV DNA test alone or combined to screening glandular lesions may contribute to minimize the limitations of both conventional and LBC preparations to diagnose glandular abnormalities.

Lymphatic vessel density and epithelial D2-40 immunoreactivity in pre-invasive and invasive lesions of the uterine cervix.

OBJECTIVE: We sought to determine the significance of lymphatic vessel density (LVD) in pre-malignant lesions and carcinomas of the uterine cervix and to evaluate the prognostic value of lymphatic invasion and D2-40 positivity in tumor cells in the three histological types of invasive lesions. The correlation of LVD, lymphatic invasion and D2-40 positivity in tumor cells with EGFR and COX-2 expressions was also evaluated. METHODS: We studied 50 cervicitis, 50 low-grade squamous intraepithelial lesions (LSIL) (CIN1), 51 high-grade squamous intraepithelial lesions (HSIL) (CIN2/CIN3), 49 invasive squamous cells carcinomas (SCC), 43 adenocarcinomas (AC) and 30 adenosquamous cells carcinomas (ASC). The immunoreaction assay was performed using the monoclonal antibody D2-40. RESULTS: Significant differences in LVD were found among all categories of pre-invasive and invasive lesions (p=0.001 and p<0.001, respectively). LVD in invasive lesions was significantly greater than in pre-invasive lesions (p<0.001) and no significant association was found between LVD in invasive lesions and both lymph node invasion and/or metastasis. D2-40 positivity in tumor cells was associated with a better prognosis in ASC cases. EGFR and COX-2 expressions in invasive lesions were not associated with LVD; however, they correlated with both lymphatic invasion and D2-40 positivity in tumor cells. CONCLUSIONS: Lymphatic neovascularization begins early in intraepithelial lesions and continues to increase towards malignancy. Both lymphatic invasion and decrease in D2-40 expression in tumor cells appear to have a prognostic value.

Cyclooxygenase-2 and epidermal growth factor receptor expressions in different histological subtypes of cervical carcinomas.

This study was designed to evaluate the significance of cyclooxygenase (COX)-2 and epidermal growth factor receptor (EGFR) expression in a series of cervical adenocarcinoma (AC), cervical adenosquamous carcinoma (ASC), and cervical squamous cell carcinoma (SCC). One hundred thirty cases of cervical carcinoma (30 ASC, 50 AC, and 50 SCC) were analyzed for COX-2 and EGFR expressions using specific primary antibodies. Samples were scored semiquantitatively as follows: (-), 0% of immunoreactive cells; (+), <5% of immunoreactive cells; (++), 5% to 50% of immunoreactive cells; and (+++), >50% of immunoreactive cells. The COX-2 expression was more frequently positive than EGFR in all cervical cancers studied. The COX-2 expression was also more prominent in AC than in ASC (P = 0.003). Expression of either COX-2 or EGFR was significantly different when comparing SCC with AC (P < 0.001 and P = 0.04, respectively). There was no significant correlation between COX-2 and EGFR expressions and age at diagnosis, recurrence, distant metastasis, and/or positive status of regional lymph nodes, neither between COX-2 and EGFR coexpression and the clinical data analyzed. Nevertheless, our data support that there are significant differences between EGFR and COX-2 expressions in the 3 different histogenetic types of cervical cancer. Also, in terms of therapeutic strategies, our data can be valuable in the selection of patients eligible to receive specific EGFR/COX-2-targeted therapy.

Toxoplasmose pulmonar associada a criptococose no SNC / Pulmonary toxoplasmosis associated to cryptococcosis of CNS

Toxoplasmose pulmonar foi diagnosticada pela lavado broncoalveolar em uma paciente de 34 anos, HIV positiva, com lesäo cavitária isolada em pulmäo direito. Criptococose foi simultaneamente diagnosticada pela cultura do líquor. O comprometimento pulmonar pelo T. gondii é discutido, bem como a associaçäo com criptococose do SNC

Prevalência do "Helicobacter pylory" na hipertençäo portal / Incidence of \"Helicobacter pylory\" in portal hypertension

Estudou-se a prevalência do Helicobacter pylori (Hp) em 50 pacientes com hipertensao portal (HP) de origem cirrótica (26), esquistossomótica (17) e outras causas (7) e sua posível relaçao com episódios prévios de hemorragia alta (HDA) e gastropatia congestiva (GC). Cem pacientes com dispepsia nao-ulcerosa constituíram o grupo controle. O Hp foi positivo em 81 por cento nos controles e em 56 por cento do grupo com HP (P<0,05) nao houve diferença significativa em relaçao à presença do Hp nas diferentes etiologias da HP. Foram positivos para o Hp 62,5 por cento dos pacientes com sangramento prévio de 44,4 por cento dos sem sangramento (p>0,05-NS). Cinqüenta por cento dos pacientes com GC e 66,6 por cento dos sem GC foram positivos para o Hp (p>0,05-NS). Nossos resultados sugerem que o Hp nao se relaciona com a HDA da HP nem com a GC; dessa forma, nao deve estar envolvido com as complicaçoes da síndrome. Acreditamos que nao há indicaçao para o tratamento do Hp como forma de prevenir ou atenuar o risco de sangramento na HP.