Cancers Of Epithelial Origin Research Articles

CURRENT OPINIONS CONCERNING THE PATHOGENESIS OF EPITHELIAL OVARIAN CANCER AND NEW DIAGNOSTIC APPROACHES TO THE DISEASE

Because of high mortality rate, the lack of effective methods for early diagnosis and limited success in treatment, ovarian cancer of epithelial origin is a major problem in gynecological oncology. These problems are largely due to our lack of the disease etiopathogenesis understanding. This article attempts to summarize recent data on the sources and the development of the cancer process in the ovary as well as to introduce modern approaches to the diagnosis of this type of malignancy.

EGFR-Signaling and Autophagy: How they Fit in the Cancer Landscape

Key significance of an overachieving Epidermal Growth Factor Receptor (EGFR)-signaling in cancer aggressiveness and poor prognosis is well recognized. In accordance, EGFR is either amplified or mutated in majority of the cancers of epithelial origin, and therefore has been recognized as a principal target for anticancer therapy. However, despite initial clinical efficacy of the anti-EGFR therapy in cancer treatment, long-term attempt to mute the cancer boosting effects of EGFR-dependent signaling meets resistance in cancer cells. Notably, effects of EGFR activation are pleotropic. Also, under conditions of anti-EGFR therapy in cancer cells, feedback activation of the pro-survival signaling by activation of other growth factor receptors can occur. However, a critical role of autophagy in the resistance against anti-EGFR therapy is fast emerging. Interestingly, EGFR regulates autophagy in a context-dependent manner. Furthermore, EGFR deregulated tumors demonstrate differential dependence upon autophagy for their survival and growth. Also, inhibiting EGFR-signaling promotes autophagy. These intriguing considerations are complicated further by findings that EGFR regulates autophagy in kinase-dependent or independent manner. Thus, for effective clinical cancer treatment using anti-EGFR regimen, it is critical that we understand molecular details of the nexus between the EGFR-signaling and autophagy.

EPITHELIAL OVARIAN CANCER. CLINICAL LECTURE

About 90 percent of ovarian cancers are epithelial tumors. High mortality rates, the lack of effective methods for early diagnosis and treatment make of the ovarian cancer a relevant problem for modern oncology. Only a wide application of clinical guidelines developed by professional societies based on the principles of evidence-based medicine could improve prognosis for that group of patients. This article attempts to summarize the experience of the world's leading centers for the fight against ovarian cancer of epithelial origin and offer solutions relevant to everyday clinical practice.

Triple synchronous primary gynaecological tumours. A case report

Abstract Background Synchronous multiple primary malignancies in the female genital tract are infrequent. From 50 to 70% of them corresponds to synchronous cancers of the endometrium and ovary. To our knowledge, this is only the third case report in the international literature of three concurrent gynaecological cancers of epithelial origin. A case is presented, as well as a literature review due to the infrequency of its diagnosis and the lack of information on the subject. Clinical case A 49-year-old woman, with previous gynaecological history of ovarian endometriosis. She underwent a hysterectomy and bilateral oophorectomy, as she had been diagnosed with endometrial hyperplasia with atypia. The final histopathology reported synchronous ovarian, Fallopian tube, and endometrial cancer. An extension study and complete surgical staging was performed, both being negative. She received adjuvant treatment of chemotherapy and radiotherapy. She is currently free of disease. Conclusions The aetiology is uncertain. There is controversy relating to increased susceptibility of synchronous neoplasms to pelvic endometriosis and inherited genetic syndromes. Its diagnosis needs to differentiate them from metastatic disease. Additionally, they are problematical from a clinical, diagnostic, therapeutic, and prognostic point of view. The presentation of more cases of triple synchronous cancers is necessary for a complete adjuvant and surgical treatment.

Using aptamers to elucidate esophageal cancer clinical samples.

The epithelial cell adhesion molecule (EpCAM) is closely correlated with the occurrence and development of various cancers of epithelial origin. This study tested, for the first time, the ability of EpCAM aptamer SYL3C to detect EpCAM expression in 170 cases of esophageal cancer (EC) and precancerous lesions, as well as 20 cases of EC series samples, using immunofluorescence imaging analysis. Corresponding antibodies were used as control. EpCAM overexpression was 98% in both esophageal squamous cell carcinoma (ESCC) and esophageal adenocarcinoma (EACA) and 100% in metastasis, but no EpCAM overexpression was detected in undifferentiated EC (UEC). Significant differences were noted among various stages of differentiation (p < 0.05) with the degree of differentiation inversely correlated with the expression of EpCAM. Overexpressed EpCAM was detected in severe dysplasia, but negative in mild to moderate dysplasia and benign esophageal lesions. In a competitive binding experiment, EpCAM aptamer generated a staining pattern similar to that of antibody, but the binding sites with EpCAM were different. Based on these results, it can be concluded that EpCAM is suitable for use as an EC biomarker, therapeutic target, and effective parameter for tumor transfer and prognosis evaluation by aptamer SYL3C staining.

Surface engineered magnetic nanoparticles for specific immunotargeting of cadherin expressing cells

In spite of historic advances in cancer biology and recent development of sophisticated chemotherapeutics, the outlook for patients with advanced cancer is still grim. In this sense nanoparticles (NPs), through their unique physical properties, enable the development of new approaches for cancer diagnosis and treatment. Thus far the most used active targeting scheme involves NPs functionalization with antibodies specific to molecules overexpressed on cancer cell’s surface. Therefore, such active targeting relies on differences in NPs uptake kinetics rates between tumor and healthy cells. Many cancers of epithelial origin are associated with the inappropriate expression of non-epithelial cadherins (e.g. N-, P-, -11) with concomitant loss of E-cadherin. Such phenomenon named cadherin switching favors tumor development and metastasis via interactions of tumor cells with stromal components. That is why we optimized the oriented functionalization of fluorescently labelled magnetic NPs with a novel antibody specific for the extracellular domain of cadherin-11. The obtained Ab-NPs exhibited high specificity when incubated with two cell lines used as models of tumor and healthy cells. Thus, cadherin switching offers a great opportunity for the development of active targeting strategies aimed to improve the early detection and treatment of cancer.

EBV BART MicroRNAs Target Multiple Pro-apoptotic Cellular Genes to Promote Epithelial Cell Survival.

Epstein-Barr virus (EBV) is a ubiquitous human γ-herpesvirus that can give rise to cancers of both B-cell and epithelial cell origin. In EBV-induced cancers of epithelial origin, including nasopharyngeal carcinomas (NPCs) and gastric carcinomas, the latent EBV genome expresses very high levels of a cluster of 22 viral pre-miRNAs, called the miR-BARTs, and these have previously been shown to confer a degree of resistance to pro-apoptotic drugs. Here, we present an analysis of the ability of individual miR-BART pre-miRNAs to confer an anti-apoptotic phenotype and report that five of the 22 miR-BARTs demonstrate this ability. We next used photoactivatable ribonucleoside-enhanced crosslinking and immunoprecipitation (PAR-CLIP) to globally identify the mRNA targets bound by these miR-BARTs in latently infected epithelial cells. This led to the identification of ten mRNAs encoding pro-apoptotic mRNA targets, all of which could be confirmed as valid targets for the five anti-apoptotic miR-BARTs by indicator assays and by demonstrating that ectopic expression of physiological levels of the relevant miR-BART in the epithelial cell line AGS resulted in a significant repression of the target mRNA as well as the encoded protein product. Using RNA interference, we further demonstrated that knockdown of at least seven of these cellular miR-BART target transcripts phenocopies the anti-apoptotic activity seen upon expression of the relevant EBV miR-BART miRNA. Together, these observations validate previously published reports arguing that the miR-BARTs can exert an anti-apoptotic effect in EBV-infected epithelial cells and provide a mechanistic explanation for this activity. Moreover, these results identify and validate a substantial number of novel mRNA targets for the anti-apoptotic miR-BARTs.

Abstract P6-01-15: Expression of high affinity folate receptor in breast cancer brain metastasis

Abstract Background: Folic acid is required by proliferating cells for the synthesis of nucleotide bases among other tasks. The high affinity folate receptor (HFR) is a membrane protein that is upregulated in several cancers of epithelial origin and rarely present in most normal cells. High expression of HFR has been documented in ovarian, breast, kidney, uterine and lung cancers. In breast cancer patients, HFR is overexpressed in 33% of primary tumors (PT) and this is associated with poor prognosis. The HFR expression in breast cancer brain metastases (BCBM) is unknown. The aim of this study was to analyze the incidence of HFR expression in BCBM and its role in the prognosis of this high-risk patient subgroup. Methods: We analyzed a database of 42 patients who underwent craniotomy with resection of breast cancer brain metastasis (BCBM). Of those, we collected 19 brain metastasis (BM) and 13 PT. HFR status was assessed by immunohistochemistry. Log-Rank test analyzed differences in overall survival (OS) between groups. Results: Median age was 51 years (range 24-74). Median follow-up was 4.2 years (range 0.6-18.5). HFR was positive in 4/19 BM (21.1%) and in 1/13 PT (7.7%). Positive samples had low H-scores (range 1-50). Staining with H&amp;E revealed that 11/19 BM (57.9%) and 7/13 PT (53.8%) had apocrine differentiation. Analysis of OS showed no difference between patients with positive HFR (median OS 48 months) and negative HFR (median OS 69 months) (P=0.25). Similarly, there was no difference in OS between patients with apocrine differentiation (median OS 63 months) and those without apocrine differentiation (median OS 69 months) (P=0.49). Conclusions: To the best of our knowledge, this is the first analysis of HFR expression in BCBM. While previous studies associated the presence of HFR with worse prognosis; in this cohort of high-risk patients, HFR was positive in only 21.1% of BM with low levels of positivity. Around half of our patients had apocrine differentiation. Neither the status of HFR nor the presence of apocrine features had impact in OS in our cohort. Citation Format: Jose P Leone, Rohit Bhargava, Adrian Lee, Adam Brufsky. Expression of high affinity folate receptor in breast cancer brain metastasis [abstract]. In: Proceedings of the Thirty-Seventh Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2014 Dec 9-13; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2015;75(9 Suppl):Abstract nr P6-01-15.

Using DNA aptamer probe for immunostaining of cancer frozen tissues.

Tissue immunostaining is critically important in clinical applications, and antibodies have been used extensively as the molecular probes. Recently, aptamer, as a new class of probes, have attracted much attention for their potential clinical and research value. Epithelial cell adhesion molecule (EpCAM) is a specific biomarker which is overexpressed in many cancers of epithelial origin. Here, a DNA-based EpCAM aptamer SYL3C is reported as a probe for the immunostaining of frozen and paraffin-embedded sections of colorectal cancer tissues. Commercialized EpCAM antibodies were also used as a standard control. EpCAM aptamer SYL3C specifically recognized and immunostained cancer nests of colorectal tumor sections, but it neither reacted with background cells within tumor sites nor exhibited cross-reaction to the benign lesions or inflammation of colorectal tissues. No cross-linking to EpCAM-negative malignant tumor sections occurred. Compared with standard antibody staining, our EpCAM aptamer SYL3C protocol is simpler to implement with a shorter reaction time. Moreover, SYL3C can specifically bind with either frozen or paraffin-embedded tissue sections. Since the histopathology of frozen tissue is closer to that of fresh tissue and since frozen sections can be produced more quickly than paraffin-embedded sections, SYL3C immunostaining of frozen sections is a quick protocol that is easy to implement.

Analysis of Antifolate Drugs with Disease Tissue Specificity

Antifolates, analogues of the essential vitamin folic acid, are used in the clinic to treat cancers and inflammatory diseases. Antifolates are primarily transported into cells via the endogenously expressed reduced folate carrier (RFC). Conversely, our collaborators in Aleem Gangjee's group at Duquesne University have synthesized antifolates (AG antifolates) that transport poorly by the RFC, but are efficiently transported by the folate receptor (hFR). The GPI-anchored hFR is lowly expressed on the apical surface in a subset of normal epithelial lineages, but is highly expressed in many cancers of epithelial origin and on activated macrophages in inflammatory disease. Therefore, AG antifolate molecules have specificity for transport into disease cells over healthy cells. These newly developed AG antifolates cause cell death via inhibition of an enzyme involved in de novo purine synthesis, glycinamide ribonucleotide (GAR) transformylase.We analyzed a series of AG antifolates using biophysical and biochemical techniques to understand both the specificity for transport by the folate receptor as well as the inhibition of the GAR transformylase in order to drive informed, hypothesis-based drug design. Our data, including pH-dependent binding profiles, enzyme inhibition data, and crystallographic models of protein in complex with AG molecules will be presented in the context of drug design and development.

Protease‐activated receptor‐1 drives pancreatic cancer progression and chemoresistance

Protease activated receptor (PAR)-1 expression in tumor cells is associated with disease progression and overall survival in a variety of cancers of epithelial origin; however, the importance of PAR-1 in the tumor microenvironment remains unexplored. Utilizing an orthotopic pancreatic cancer model in which tumor cells are PAR-1 positive whereas stromal cells are PAR-1 negative, we show that PAR-1 expression in the microenvironment drives progression and induces chemoresistance of pancreatic cancer. PAR-1 enhances monocyte recruitment into the tumor microenvironment by regulating monocyte migration and fibroblast dependent chemokine production thereby inducing chemoresistance. Overall, our data identify a novel role of PAR-1 in the pancreatic tumor microenvironment and suggest that PAR-1 may be an attractive target to reduce drug resistance in pancreatic cancer.

The Role and Clinical Relevance of Disseminated Tumor Cells in Breast Cancer

Tumor cell dissemination is a common phenomenon observed in most cancers of epithelial origin. One-third of breast cancer patients present with disseminated tumor cells (DTCs) in bone marrow at time of diagnosis; these patients, as well as patients with persistent DTCs, have significantly worse clinical outcome than DTC-negative patients. Since DTC phenotype may differ from the primary tumor with regard to ER and HER2 status, reevaluation of predictive markers on DTCs may optimize treatment choices. In the present review, we report on the clinical relevance of DTC detection in breast cancer.

ERM proteins in cancer progression

Members of the ezrin-radixin-moesin (ERM) family of proteins are involved in multiple aspects of cell migration by acting both as crosslinkers between the membrane, receptors and the actin cytoskeleton, and as regulators of signalling molecules that are implicated in cell adhesion, cell polarity and migration. Increasing evidence suggests that the regulation of cell signalling and the cytoskeleton by ERM proteins is crucial during cancer progression. Thus, both their expression levels and subcellular localisation would affect tumour progression. High expression of ERM proteins has been shown in a variety of cancers. Mislocalisation of ERM proteins reduces the ability of cells to form cell-cell contacts and, therefore, promotes an invasive phenotype. Similarly, mislocalisation of ERM proteins impairs the formation of receptor complexes and alters the transmission of signals in response to growth factors, thereby facilitating tumour progression. In this Commentary, we address the structure, function and regulation of ERM proteins under normal physiological conditions as well as in cancer progression, with particular emphasis on cancers of epithelial origin, such as those from breast, lung and prostate. We also discuss any recent developments that have added to the understanding of the underlying molecular mechanisms and signalling pathways these proteins are involved in during cancer progression.

Pathway Modulations and Epigenetic Alterations in Ovarian Tumorbiogenesis

Cellular pathways are numerous and are highly integrated in function in the control of cellular systems. They collectively regulate cell division, proliferation, survival and apoptosis of cells and mutagenesis of key genes that control these pathways can initiate neoplastic transformations. Understanding these pathways is crucial to future therapeutic and preventive strategies of the disease. Ovarian cancers are of three major types; epithelial, germ-cell, and stromal. However, ovarian cancers of epithelial origin, arising from the mesothelium, are the predominant form. Of the subtypes of ovarian cancer, the high-grade serous tumors are fatal, with low survival rate due to late detection and poor response to treatments. Close examination of preserved ovarian tissues and in vitro studies have provided insights into the mechanistic changes occurring in cells mediated by a few key genes. This review will focus on pathways and key genes of the pathways that are mutated or have aberrant functions in the pathology of ovarian cancer. Non-genetic mechanisms that are gaining prominence in the pathology of ovarian cancer, miRNAs and epigenetics, will also be discussed in the review.

Triple simultaneous primary invasive gynecological malignancies: A case report

Double gynecologic cancer (primary cancers in two organs) is relatively rare. However, triple gynecologic cancer (primary cancers in three organs) is extremely rare. We experienced a case of triple cancer, with primary cervical, endometrial and ovarian cancers, each showing different histopathological features. A 50-year-old woman with a preoperative diagnosis of cervical cancer stage Ib1 with a pathological diagnosis of mucinous adenocarcinoma underwent radical hysterectomy. The pathological diagnoses of the extracted masses were endometrioid adenocarcinoma in the uterine corpus and serous adenocarcinoma in the left ovary. Consequently, triple cancer was diagnosed. After the operation, six cycles of a paclitaxel/carboplatin regimen were administered, and no relapse of the cancers has been observed to date. To our knowledge, this is only the second case report in the international literature of concurrent gynecologic triple cancers of epithelial origin; that is, invasive cervical, endometrial and ovarian cancers, each with different histopathological features.

EYA4 is inactivated biallelically at a high frequency in sporadic lung cancer and is associated with familial lung cancer risk

In an effort to identify novel biallelically inactivated tumor suppressor genes (TSG) in sporadic invasive and pre-invasive non-small cell lung cancer (NSCLC) genomes, we applied a comprehensive integrated multi-‘omics approach to investigate patient matched, paired NSCLC tumor and non-malignant parenchymal tissues. By surveying lung tumor genomes for genes concomitantly inactivated within individual tumors by multiple mechanisms, and by the frequency of disruption in tumors across multiple cohorts, we have identified a putative lung cancer TSG, Eyes Absent 4 (EYA4). EYA4 is frequently and concomitantly deleted, hypermethylated and underexpressed in multiple independent lung tumor data sets, in both major NSCLC subtypes, and in the earliest stages of lung cancer. We find not only that decreased EYA4 expression is associated with poor survival in sporadic lung cancers, but EYA4 SNPs are associated with increased familial cancer risk, consistent with EYA4’s proximity to the previously reported lung cancer susceptibility locus on 6q. Functionally, we find that EYA4 displays TSG-like properties with a role in modulating apoptosis and DNA repair. Cross examination of EYA4 expression across multiple tumor types suggests a cell type-specific tumorigenic role for EYA4, consistent with a tumor suppressor function in cancers of epithelial origin. This work shows a clear role for EYA4 as a putative TSG in NSCLC.

Knockdown of core binding factorβ alters sphingolipid metabolism

Core binding factor (CBF) is a heterodimeric transcription factor containing one of three DNA-binding proteins of the Runt-related transcription factor family (RUNX1-3) and the non-DNA-binding protein, CBFβ. RUNX1 and CBFβ are the most common targets of chromosomal rearrangements in leukemia. CBF has been implicated in other cancer types; for example RUNX1 and RUNX2 are implicated in cancers of epithelial origin, including prostate, breast, and ovarian cancers. In these tumors, CBF is involved in maintaining the malignant phenotype and, when highly over-expressed, contributes to metastatic growth in bone. Herein, lentiviral delivery of CBFβ-specific shRNAs was used to achieve a 95% reduction of CBFβ in an ovarian cancer cell line. This drastic reduction in CBFβ expression resulted in growth inhibition that was not associated with a cell cycle block or an increase in apoptosis. However, CBFβ silencing resulted in increased autophagy and production of reactive oxygen species (ROS). Since sphingolipid and ceramide metabolism regulates non-apoptotic cell death, autophagy, and ROS production, fumonsin B1 (FB1), an inhibitor of ceramide synthase, was used to alter ceramide production in the CBFβ-silenced cells. FB1 treatment inhibited the CBFβ-dependent increase in autophagy and provided a modest increase in cell survival. To document alterations to sphingolipids in the CBFβ-silenced cells, ceramide, and lactosylceramide levels were directly examined by mass spectrometry. Substantial increases in ceramide species and decreases in lactosylceramides were identified. Altogether, this report provides evidence that CBF transcriptional pathways control cellular survival, at least in part, through sphingolipid metabolism.

Abstract 331: Head and neck and skin squamous cell carcinoma developed in a BRCA2 knockout mouse model.

Abstract BRCA1 and BRCA2 were originally identified as breast cancer tumor suppressor genes. Germline mutations of BRCA1 have been found to predispose the carrier mainly to breast and ovarian cancers. However, germline mutations of BRCA2 can cause a much wider spectrum of cancers, including breast, ovarian, pancreatic and prostate cancers. There are conflicting reports on whether BRCA2 mutations also contribute to the tumorigenesis of head and neck squamous cell carcinoma (HNSCC). Previously, it had been reported that K14-cre; BRCA2f/f mice do not develop any cancer of epithelial origin even at 600 days of age in a 129/C57 background. Here, we generated a K14-cre mediated BRCA2 knockout mouse model in a different mouse strain background than previously reported. To our surprise, we found that K14-cre; BRCA2 f/f mice started to develop tumors at seven months of age. Most of the tumors were in the head and neck region, while others were skin tumors in other locations. The head and neck tumors showed no skin involvement or ulceration. Tumor growth in many mice eventually affected their food intake and they had to be euthanized for humane reason. At around 11 months of age, we sacrificed the remaining mice and subjected them to necroscopy. All the visually suspicious tissues were processed for pathologic examination. In total, we found 8 out of 31 K14-cre; BRCA2 f/f mice (26%) with pathologically confirmed head and neck cancer and 4 out 31 K14-cre; BRCA2 f/f mice (13%) with skin cancer. None of the 28 control BRCA2 f/f mice developed any cancer at this age. The head and neck cancers are squamous cell carcinomas arising from the mucosa surface and form large, palpable masses. In some areas, the tumors are well-differentiated with evidence of abundant keratin production, while in other areas they are less differentiated and exhibit high nuclear to cytoplasmic ratios, nuclear aytpia, mitoses and central necrosis. These histological features are highly reminiscent of the pathological characteristics of human HNSCC. This finding strongly indicates that mutations of BRCA2 play an important role in the tumorigenesis of HNSCC in a specific genetic background. Citation Format: Michael Kamradt, Chaozhong Zou, Lin Liu, Yuexia Wang, Philip P. Fitchev, Howard J. Worman, Susan E. Crawford, Mark Talamonti, Qingshen Gao. Head and neck and skin squamous cell carcinoma developed in a BRCA2 knockout mouse model. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 331. doi:10.1158/1538-7445.AM2013-331

EMT in developmental morphogenesis

Carcinomas, cancers of epithelial origin, constitute the majority of all cancers. Loss of epithelial characteristics is an early step in carcinoma progression. Malignant transformation and metastasis involve additional loss of cell-cycle control and gain of migratory behaviors. Understanding the relationships among epithelial homeostasis, cell proliferation, and cell migration is therefore fundamental in understanding cancer. Interestingly, these cellular events also occur frequently during animal development, but without leading to tumor formation. Can we learn anything about carcinomas from developmental biology? In this review, we focus on one aspect of carcinoma progression, the Epithelial–Mesenchymal Transition (EMT), and provide an overview of how the EMT is involved in normal amniote development. We discuss 12 developmental and morphogenetic processes that clearly involve the EMT. We conclude by emphasizing the diversity of EMT processes both in terms of their developmental context and of their cellular morphogenesis. We propose that there is comparable diversity in cancer microenvironment and molecular regulation of cancer EMTs.

SOX4 Mediates TGF-β-Induced Expression of Mesenchymal Markers during Mammary Cell Epithelial to Mesenchymal Transition

The epithelial to mensenchymal transition program regulates various aspects of embryonic development and tissue homeostasis, but aberrant activation of this pathway in cancer contributes to tumor progression and metastasis. TGF-β potently induces an epithelial to mensenchymal transition in cancers of epithelial origin by inducing transcriptional changes mediated by several key transcription factors. Here, we identify the developmental transcription factor SOX4 as a transcriptional target of TGF-β in immortalized human mammary epithelial cells. SOX4 expression and activity are rapidly induced in the early stages of the TGF-β-induced epithelial to mensenchymal transition. We demonstrate that conditional activation of Sox4 is sufficient to induce the expression of N-cadherin and additional mesenchymal markers including vimentin and fibronectin, but fails to induce complete EMT as no changes are observed in the expression of E-cadherin and β-catenin. Moreover, shRNA-mediated knockdown of SOX4 significantly delays TGF-β-induced mRNA and protein expression of mesenchymal markers. Taken together, these data suggest that TGF-β-mediated increased expression of SOX4 is required for the induction of a mesenchymal phenotype during EMT in human mammary epithelial cells.