Evolution of intra-tumoral heterogeneity across different pathological stages in papillary thyroid carcinoma.

BACKGROUND: Intra-tumor heterogeneity (ITH) results from the continuous accumulation of mutations during disease progression, thus impacting patients' clinical outcome. How the ITH evolves across papillary thyroid carcinoma (PTC) different tumor stages is lacking. METHODS: We used the whole-exome sequencing data from The Cancer Genome Atlas Thyroid Cancer (TCGA-THCA) cohort to track the ITH and assessed its relationship with clinical features through different stages of the PTC progression. We further assayed the expression levels of the specific genes in papillary thyroid cancer cell lines compared to an immortalized normal thyroid epithelial cell line by qRT-PCR. RESULTS: We revealed the timing of mutational processes and the dynamics of the temporal acquisition of somatic events during the lifetime of the PTC. ITH significantly influences the PTC patient's survival rate and, as genetic heterogeneity increases, the prognosis gets worse in advanced tumor stages. ITH also affects the mutational architecture of each clinical stage which is subject to periodic fluctuations. Different mutational processes may cooperate to shape a stage-specific mutational spectrum during the progression from early to advanced tumor stages. Moreover, different evolutionary paths characterize PTC progression across pathological stages due to both mutations recurrently occurring in all stages in hotspot positions and distinct codon changes dominating in different stages. A different expression level of specific genes also exists in different thyroid cancer cell lines. CONCLUSIONS: Our findings suggest ITH as a potential unfavorable prognostic factor in PTC and highlight the dynamic changes in different clinical stages of PTC, providing some clues for the precision medicine and suggesting different diagnostic decisions depending on the clinical stages of patients. Finally, complete clear guidelines to define risk stratification of PTC patients are lacking; thus, this work could contribute to defining patients who need more aggressive treatments and, in turn, could reduce the social burden of this cancer.

miR-622 is a novel potential biomarker of breast carcinoma and impairs motility of breast cancer cells through targeting NUAK1 kinase.

BACKGROUND: Aberrant expression of microRNAs (miR) has been proposed as non-invasive biomarkers for breast cancers. The aim of this study was to analyse the miR-622 level in the plasma and in tissues of breast cancer patients and to explore the role of miR-622 and its target, the NUAK1 kinase, in this context. METHODS: miR-622 expression was analysed in plasma and in tissues samples of breast cancer patients by q-RT-PCR. Bioinformatics programs, luciferase assay, public dataset analysis and functional experiments were used to uncover the role of miR-622 and its target in breast cancer cells. RESULTS: miR-622 is downregulated in plasma and in tissues of breast cancer patients respect to healthy controls and its downregulation is significantly associated with advanced grade and high Ki67 level. Modulation of miR-622 affects the motility phenotype of breast cancer cells. NUAK1 kinase is a functional target of miR-622, it is associated with poor clinical outcomes of breast cancer patients and is inversely correlated with miR-622 level. CONCLUSIONS: miR-622/NUAK1 axis is deregulated in breast cancer patients and affects the motility phenotype of breast cancer cells. Importantly, miR-622 and NUAK1 hold promises as biomarkers and as targets for breast cancers.

The TUSC2 Tumour Suppressor Inhibits the Malignant Phenotype of Human Thyroid Cancer Cells via SMAC/DIABLO Protein.

Thyroid carcinoma is the most common endocrine cancer and includes different forms. Among these, anaplastic thyroid carcinoma (ATC) is the rarest but the most lethal subtype, compared to papillary thyroid carcinoma (PTC) which shows an overall good prognosis. We have previously showed that Tumor Suppressor Candidate 2 (TUSC2), a known tumour suppressor gene, is downregulated in human PTC and ATC compared to normal thyroid samples. The aim of this study was to gain insight into the molecular mechanisms induced by TUSC2 in thyroid cancer cells. Here, we stably transfected TUSC2 in papillary (TPC-1) and in anaplastic (8505C) thyroid cancer cell lines and studied its effects on several biological processes, demonstrating that TUSC2 overexpression decreased thyroid cancer cell proliferation, migration and invasion. Through the proteome profiler apoptosis array, we observed that TUSC2 increased sensitivity to apoptosis by increasing the SMAC/DIABLO and CYTOCHROME C proteins. On the other hand, transient silencing of TUSC2, by siRNA, in an immortalized thyroid follicular epithelial cell line (Nthy-ori 3-1) showed the opposite effect. Finally modulation of SMAC/DIABLO partially rescued the biological effects of TUSC2. Thus, our data highlight a tumour suppressor role of TUSC2 in thyroid carcinogenesis, suggesting that it could be a promising target and biomarker for thyroid carcinoma.

Junctional adhesion molecule-A is down-regulated in anaplastic thyroid carcinomas and reduces cancer cell aggressiveness by modulating p53 and GSK3 α/ß pathways.

Junctional adhesion molecule A (JAM-A) is a transmembrane protein that contributes to different biological process, including the epithelial to mesenchymal transition (EMT). Through an EMT profiler array, we explored the molecular players associated with human thyroid cancer progression and identified JAM-A as one of the genes mostly deregulated. The quantitative real-time polymerase chain reaction and immunohistochemistry analyses showed that downregulation of JAM-A occurred in anaplastic thyroid carcinoma (ATC) compared with normal thyroid (NT) and papillary thyroid carcinoma (PTC) tissues and correlated with extrathyroid infiltration, tumor size, and ATC histotype. In ATC cell lines, JAM-A restoration suppressed malignant hallmarks of transformation including cell proliferation, motility, and transendothelial migration. Accordingly, knockdown of JAM-A enhanced thyroid cancer cell proliferation and motility in PTC cells. Through the proteome profiler human phospho-kinase array, we demonstrated that higher expression of JAM-A was associated with a significant increased level of phosphorylation of p53 and GSK3 α/ß proteins. In conclusion, our findings highlight a novel role of JAM-A in thyroid cancer progression and suggest that JAM-A restoration could have potential clinical relevance in thyroid cancer treatment.

Multimodal imaging for a theranostic approach in a murine model of B-cell lymphoma with engineered nanoparticles.

Nanoparticles (NPs) are a promising tool for in vivo multimodality imaging and theranostic applications. Hyaluronic acid (HA)-based NPs have numerous active groups that make them ideal as tumor-targeted carriers. The B-lymphoma neoplastic cells express on their surfaces a clone-specific immunoglobulin receptor (Ig-BCR). The peptide A20-36 (pA20-36) selectively binds to the Ig-BCR of A20 lymphoma cells. In this work, we demonstrated the ability of core-shell chitosan-HA-NPs decorated with pA20-36 to specifically target A20 cells and reduce the tumor burden in a murine xenograft model. We monitored tumor growth using high-frequency ultrasonography and demonstrated targeting specificity and kinetics of the NPs via in vivo fluorescent reflectance imaging. This result was also confirmed by ex vivo magnetic resonance imaging and confocal microscopy. In conclusion, we demonstrated the ability of NPs loaded with fluorescent and paramagnetic tracers to act as multimodal imaging contrast agents and hence as a non-toxic, highly specific theranostic system.

Preclinical Imaging for the Study of Mouse Models of Thyroid Cancer.

Thyroid cancer, which represents the most common tumors among endocrine malignancies, comprises a wide range of neoplasms with different clinical aggressiveness. One of the most important challenges in research is to identify mouse models that most closely resemble human pathology; other goals include finding a way to detect markers of disease that common to humans and mice and to identify the most appropriate and least invasive therapeutic strategies for specific tumor types. Preclinical thyroid imaging includes a wide range of techniques that allow for morphological and functional characterization of thyroid disease as well as targeting and in most cases, this imaging allows quantitative analysis of the molecular pattern of the thyroid cancer. The aim of this review paper is to provide an overview of all of the imaging techniques used to date both for diagnosis and theranostic purposes in mouse models of thyroid cancer.

Anterior gradient protein 2 promotes survival, migration and invasion of papillary thyroid carcinoma cells.

BACKGROUND: Through a transcriptome microarray analysis, we have isolated Anterior gradient protein 2 (AGR2) as a gene up-regulated in papillary thyroid carcinoma (PTC). AGR2 is a disulfide isomerase over-expressed in several human carcinomas and recently linked to endoplasmic reticulum (ER) stress. Here, we analyzed the expression of AGR2 in PTC and its functional role. METHODS: Expression of AGR2 was studied by immunohistochemistry and real time PCR in normal thyroids and in PTC samples. The function of AGR2 was studied by knockdown in PTC cells and by ectopic expression in non-transformed thyroid cells. The role of AGR2 in the ER stress was analyzed upon treatment of cells, expressing or not AGR2, with Bortezomib and analyzing by Western blot the expression levels of GADD153. RESULTS: PTC over-expressed AGR2 at mRNA and protein levels. Knockdown of AGR2 in PTC cells induced apoptosis and decreased migration and invasion. Ectopic expression of AGR2 in non-transformed human thyroid cells increased migration and invasion and protected cells from ER stress induced by Bortezomib. CONCLUSIONS: AGR2 is a novel marker of PTC and plays a role in thyroid cancer cell survival, migration, invasion and protection from ER stress.

miR-331-5p Affects Motility of Thyroid Cancer Cell Lines and Regulates BID Expression.

During tumorigenesis, miRNAs with unbalanced expression profiles can increase the threat of disease progression. Here, we focus on the role of miR-331-5p in the pathogenesis of thyroid cancer (TC). In vitro studies were conducted using TC cell lines after the forced expression and silencing of miR-331-5p. Cell proliferation and viability were analyzed via cell counts and colorimetric assays. Cell motility was analyzed via wound healing assays, Transwell migration and invasion assays, and Matrigel Matrix assays. The putative targets of miR-331-5p were unveiled via label-free proteomic screening and then verified using Western blot and luciferase assays. Expression studies were conducted by interrogating The Cancer Genome Atlas (TCGA). We found that ectopic miR-331-5p expression reduces TC cell motility, while miR-331-5p silencing induces the opposite phenotype. Proteomic screening revealed eight putative downregulated targets of miR-331-5p, among which BID was confirmed as a direct target. TCGA data showed the downregulation of miR-331-5p and the upregulation of BID in TC tissues. In summary, deregulation of the miR-331-5p/BID axis could enhance the aggressiveness of TC cell lines, providing new insights into the mechanisms of the progression of this disease and suggesting a potential role of the component factors as possible biomarkers in TC tissues.

Imaging of thyroid tumor angiogenesis with microbubbles targeted to vascular endothelial growth factor receptor type 2 in mice.

BACKGROUND: To evaluate whether Contrast Enhanced Ultrasund (CEUS) with microbubbles (MBs) targeted to VEGFR-2 is able to characterize in vivo the VEGFR-2 expression in the tumor vasculature of a mouse model of thyroid cancer (Tg-TRK-T1). METHODS: Animal protocol was approved by Institutional committee on Laboratory Animal Care. Contrast-enhanced ultrasound imaging with MBs targeted with an anti-VEGFR-2 monoclonal antibody (UCAVEGFR-2) and isotype control antibody (UCAIgG) was performed in 7 mice with thyroid carcinoma, 5 mice with hyperplasia or benign thyroid nodules and 4 mice with normal thyroid. After ultrasonography, the tumor samples were harvested for histological examination and VEGFR-2 expression was tested by immunohistochemistry. Data were reported as median and range. Paired non parametric Wilcoxon's test and ANOVA of Kruskal-Wallis were used. The correlation between the contrast signal and the VEGFR-2 expression was assessed by the Spearman coefficient. RESULTS: The Video intensity difference (VID) caused by backscatter of the retained UCAVEGFR-2 was significantly higher in mice harboring thyroid tumors compared to mice with normal thyroids (P < 0.01) and to mice harboring benign nodules (P < 0.01). No statistically significant differences of VID were observed in the group of mice carrying benign nodules compared to mice with normal thyroids. Moreover in thyroid tumors VID of retained VEGFR-2-targeted UCA was significantly higher than that of control UCAIgG (P <0.05). Results of immunohistochemical analysis confirmed VEGFR-2 overexpression. The magnitude of the molecular ultrasonographic signal from a VEGFR-2-targeted UCA retained by tissue correlates with VEGFR-2 expression determined by immunohistochemistry (rho 0.793, P=0.0003). CONCLUSIONS: We demonstrated that CEUS with UCAVEGFR-2 might be used for in vivo non invasive detection and quantification of VEGFR-2 expression in thyroid cancer in mice, and to differentiate benign from malignant thyroid nodules.

Development of Cyclic Peptides Targeting the Epidermal Growth Factor Receptor in Mesenchymal Triple-Negative Breast Cancer Subtype.

Triple-negative breast cancer (TNBC) is an aggressive malignancy characterized by the lack of expression of estrogen and progesterone receptors and amplification of human epidermal growth factor receptor 2 (HER2). Being the Epidermal Growth Factor Receptor (EGFR) highly expressed in mesenchymal TNBC and correlated with aggressive growth behavior, it represents an ideal target for anticancer drugs. Here, we have applied the phage display for selecting two highly specific peptide ligands for targeting the EGFR overexpressed in MDA-MB-231 cells, a human TNBC cell line. Molecular docking predicted the peptide-binding affinities and sites in the extracellular domain of EGFR. The binding of the FITC-conjugated peptides to human and murine TNBC cells was validated by flow cytometry. Confocal microscopy confirmed the peptide binding specificity to EGFR-positive MDA-MB-231 tumor xenograft tissues and their co-localization with the membrane EGFR. Further, the peptide stimulation did not affect the cell cycle of TNBC cells, which is of interest for their utility for tumor targeting. Our data indicate that these novel peptides are highly specific ligands for the EGFR overexpressed in TNBC cells, and thus they could be used in conjugation with nanoparticles for tumor-targeted delivery of anticancer drugs.

Effects of Annurca Flesh Apple Polyphenols in Human Thyroid Cancer Cell Lines.

Among natural macromolecules, the polyphenol extract from Annurca flesh (AFPE) apple could play a potential therapeutic role for a large spectrum of human cancer also by exerting antioxidant properties. Thyroid cancer is a common neoplasia in women, and it is in general responsive to treatments although patients may relapse and metastasize or therapy-related side effects could occur. In this study, we explored the effects of AFPE on papillary (TPC-1) and anaplastic (CAL62) thyroid cancer cell line proliferation and viability. We found that AFPE exposure induced a reduction of cell proliferation and cell viability in dose-dependent manner. The effect was associated with the reduction of phosphorylation of Rb protein. To study the mechanisms underlying the biological effects of AFPE treatment in thyroid cancer cells, we investigated the modulation of miRNA (miR) expression. We found that AFPE treatment increased the expression of the miR-141, miR-145, miR-200a-5p, miR-425, and miR-551b-5p. Additionally, since natural polyphenols could exert their beneficial effects through the antioxidant properties, we investigated this aspect, and we found that AFPE treatment reduced the production of reactive oxygen species (ROS) in CAL62 cells. Moreover, AFPE pretreatment protects against hydrogen peroxide-induced oxidative stress in thyroid cancer cell lines. Taken together, our findings suggest that AFPE, by acting at micromolar concentration in thyroid cancer cell lines, may be considered a promising adjuvant natural agent for thyroid cancer treatment approach.

Unveiling the miRNAs responsive to physical activity/exercise training in cancer: A systematic review.

PURPOSE: Since cancer-related miRNAs expression are affected by physical activity/exercise, they represent an attractive tool to monitor this response in healthy and in cancer subjects. Here, we aim to update the literature regarding the benefit of this axis in oncology field. METHOD: We systematically questioned the literature according to PRISMA guidelines defining our questions by PICO tool and carrying out the risk of bias and quality assessment by NOS. RESULTS: Among 751 records risen from the initial search, 28 studies resulted eligible. The majority of studies regarded breast cancer, while others were initially conducted in healthy subjects and afterwards authors speculated on the relationship between exercise-modulated miRNAs and cancer. CONCLUSION: Physical activity/exercise induces beneficial effects in the cancer prevention, prognosis and treatments. Among the miRNAs modulated by physical activity/exercise, miR-21, let-7 and miR-133 families resulted the most promising in this field.

Preclinical Imaging Evaluation of miRNAs' Delivery and Effects in Breast Cancer Mouse Models: A Systematic Review.

BACKGROUND: We have conducted a systematic review focusing on the advancements in preclinical molecular imaging to study the delivery and therapeutic efficacy of miRNAs in mouse models of breast cancer. METHODS: A systematic review of English articles published in peer-reviewed journals using PubMed, EMBASE, BIOSIS™ and Scopus was performed. Search terms included breast cancer, mouse, mice, microRNA(s) and miRNA(s). RESULTS: From a total of 2073 records, our final data extraction was from 114 manuscripts. The most frequently used murine genetic background was Balb/C (46.7%). The most frequently used model was the IV metastatic model (46.8%), which was obtained via intravenous injection (68.9%) in the tail vein. Bioluminescence was the most used frequently used tool (64%), and was used as a surrogate for tumor growth for efficacy treatment or for the evaluation of tumorigenicity in miRNA-transfected cells (29.9%); for tracking, evaluation of engraftment and for response to therapy in metastatic models (50.6%). CONCLUSIONS: This review provides a systematic and focused analysis of all the information available and related to the imaging protocols with which to test miRNA therapy in an in vivo mice model of breast cancer, and has the purpose of providing an important tool to suggest the best preclinical imaging protocol based on available evidence.

Dissecting the molecular pathways involved in the effects of physical activity on breast cancers cells: A narrative review.

Epidemiologic evidence suggests that obesity and sedentary are modifiable factors strongly associated with breast cancer risk worldwide. Since breast cancer represents the most frequent malignant neoplasm and the second cause of cancer-related deaths in women worldwide, an insight into the molecular mechanisms clarifying the effects of physical activity in breast cancer cells could have important implication for changing this cancer burden. In this narrative Review article, we summarize the current knowledge, regarding the effects of adapted physical activity program, focusing on the cellular signaling pathways activated and on the molecular markers involved in breast cancer. Regular exercise training in breast cancer patients has been shown to positively affect tumor-growth and survival rate. Indeed, emerging work demonstrates that regular exercise is able to affect multiple cancer hallmarks influencing the development and progression of cancer. In conclusion, changes in the circulating insulin, adipokines and estrogen levels, inflammation and oxidative stress could represent some of the possible biological mechanisms through which exercise may influence breast cancer development and recurrence.

The lncRNA TEX41 is upregulated in pediatric B-Cells Acute Lymphoblastic Leukemia and it is necessary for leukemic cell growth.

BACKGROUND: Long non-coding RNAs (lncRNAs) represent a diverse class of RNAs involved in the regulation of various physiological and pathological cellular processes, including transcription, intracellular trafficking, and chromosome remodeling. LncRNAs deregulation was linked to the development and progression of various cancer types, such as acute leukemias. In this context, lncRNAs were also evaluated as a novel class of biomarkers for cancer diagnosis and prognosis. Here, we analyzed TEX41 in childhood B cell acute lymphoid leukemia (B-ALL). METHODS: Total RNA was extracted from pediatric B-ALL patients (at diagnosis and after induction of therapy) and from healthy subjects. Total RNA was also extracted from different leukemia cell line models. The expression level of TEX41 was evaluated by q-RT-PCR. Also, the dataset deposited by St. Jude Children's Research Hospital was consulted. Furthermore, the silencing of TEX41 in RS4;11 cell line was obtained by 2'-Deoxy, 2'Fluroarabino Nucleic Acids (2'F-ANAs) Oligonucleotides, and the effect on cell proliferation was evaluated. Cell cycle progression and its regulators were analyzed by flow cytometry and immunoblotting. RESULTS: We exploited the St Jude Cloud database and found that TEX41 is a lncRNA primarily expressed in the case of B-ALL (n = 79) while its expression levels are low/absent for T-cell ALL (n = 25) and acute myeloid leukemia (n = 38). The association of TEX41 with B-ALL was confirmed by real-time PCR assays. TEX41 disclosed increased expression levels in bone marrow from patients with B-ALL at diagnosis, while its expression levels became low or absent when retested in Bone Marrow cells of the same patient after 1 month of induction therapy. Also, silencing experiments performed on RS4;11 cells showed that TEX41 downregulation impaired in vitro leukemic cell growth determining their arrest in the G2-M phase and the deregulation of cell cycle proteins. CONCLUSIONS: Our findings highlight that TEX41 is an upregulated lncRNA in the case of B-ALL and this feature makes it a novel potential biomarker for the diagnosis of this leukemia subtype in pediatric patients. Finally, TEX41 expression seems to be critical for leukemic proliferation, indeed, silencing experiments targeting TEX41 mRNA in the RS4;11 cell line hampered in vitro cell growth and cell cycle progression, by inducing G2-M arrest as confirmed propidium iodide staining and by the upregulation of p53 and p21 proteins.

Adiponectin and leptin exert antagonizing effects on proliferation and motility of papillary thyroid cancer cell lines.

Adiponectin (Acrp30) and leptin, adipokines produced and secreted mainly by the adipose tissue, are involved in human carcinogenesis. Thyroid carcinomas are frequent endocrine cancers, and several evidences suggest that they are correlated with obesity. In this study, we first analyzed the expression levels and prognostic values of Acrp30, leptin, and their receptors in thyroid cancer cells. Then, we investigated the role of Acrp30 and leptin in proliferation, migration, and invasion. We found that Acrp30 treatment alone inhibits cell proliferation and cell viability in a time and dose-dependent manner; leptin alone does not influence thyroid cancer cells (BCPAP and K1) proliferation, but the combined treatment reverts Acrp30-induced effects on cell proliferation. Additionally, through wound healing and Matrigel Matrix invasion assays, we unveiled that Acrp30 inhibits thyroid cancer cell motility, while leptin induces the opposite effect. Importantly, in the combined treatment, Acrp30 and leptin exert antagonizing effects on papillary thyroid cancer cells' migration and invasion in both BCPAP and K1 cell lines. Highlights of these studies suggest that Acrp30 and leptin could represent therapeutic targets and biomarkers for the management of thyroid cancer.

Long-Term Recreational Football Training and Health in Aging.

This narrative review aims to critically analyze the effects of exercise on health in aging. Here we discuss the main clinical and biomolecular modifications induced by long-term recreational football training in older subjects. In particular, the effects induced by long-term recreational football training on cardiovascular, metabolic and musculo-skeletal fitness, together with the modifications in the muscle expression of hallmarks related to oxidative metabolism, DNA repair and senescence suppression pathways and protein quality control mechanisms will be provided. All these topics will be debated also in terms of preventing non-communicable metabolic diseases, in order to achieve successful aging over time.

miR-650 promotes motility of anaplastic thyroid cancer cells by targeting PPP2CA.

PURPOSE: Aberrant expression of miRNAs is crucial in several tissues tumorigenesis including thyroid. Recent studies demonstrated that miR-650 plays different role depending on the cancer type. Herein, we investigated the role of miR-650 in thyroid carcinoma. METHODS: The expression of miR-650 was analyzed in human thyroid tissues by q-RT-PCR. Anaplastic (8505C, CAL62, SW1736) and papillary (TPC-1) thyroid cancer cell lines were used to dissect the role of miR-650 on malignant hallmarks of transformation. Label-free proteomic analysis was exploited to unravel the targets of miR-650, while luciferase reporter assay and functional experiments were performed to confirm a selected target. Spearman's rank correlation test was used to assess the association between miR-650 and its target in human thyroid cancer tissues. RESULTS: miR-650 is over-expressed in anaplastic (ATC) thyroid carcinoma where it enhances cell migration and invasion. Proteomic label-free and bioinformatics analysis revealed that the serine-threonine protein phosphatase 2 catalytic subunit alpha (PPP2CA) is a target of miR-650; these finding were confirmed by luciferase assay. Restoration of PPP2CA mRNA, deprived of its 3'UTR, is able to revert the malignant phenotype induced by miR-650 in HEK-293 cells. Importantly, PPP2CA is down-regulated in ATC tissues and is inversely correlated with miR-650. CONCLUSIONS: miR-650 displayed oncogenic activity in ATC cells through targeting PPP2CA phosphatase. These results suggest that miR-650/PPP2CA axis could be modulated to interfere with motile ability of thyroid carcinoma cells.

The RET/PTC-RAS-BRAF linear signaling cascade mediates the motile and mitogenic phenotype of thyroid cancer cells.

In papillary thyroid carcinomas (PTCs), rearrangements of the RET receptor (RET/PTC) and activating mutations in the BRAF or RAS oncogenes are mutually exclusive. Here we show that the 3 proteins function along a linear oncogenic signaling cascade in which RET/PTC induces RAS-dependent BRAF activation and RAS- and BRAF-dependent ERK activation. Adoptive activation of the RET/PTC-RAS-BRAF axis induced cell proliferation and Matrigel invasion of thyroid follicular cells. Gene expression profiling revealed that the 3 oncogenes activate a common transcriptional program in thyroid cells that includes upregulation of the CXCL1 and CXCL10 chemokines, which in turn stimulate proliferation and invasion. Thus, motile and mitogenic properties are intrinsic to transformed thyroid cells and are governed by an epistatic oncogenic signaling cascade.

The heterogeneous distribution of BRAF mutation supports the independent clonal origin of distinct tumor foci in multifocal papillary thyroid carcinoma.

CONTEXT: Papillary thyroid carcinoma (PTC) is frequently multifocal. Independent PTC foci may occur either from intraglandular metastases from a single dominant tumor or as unrelated neoplastic clones. In rare cases, the simultaneous presence of PTC foci of different histopathological subtypes points to independent sites of tumor formation. OBJECTIVES: We examined the pattern of BRAF mutations in noncontiguous tumor foci and node metastases from 69 patients affected by multicentric PTC. These included 19 cases characterized by the simultaneous presence of different PTC histopathological variants. DESIGN: BRAF (exon 15) mutation was examined by PCR-single strand conformational polymorphism followed by DNA sequencing in laser-capture microdissected tissue samples. RESULTS: Discordant patterns of BRAF mutation were found in about 40% of the multifocal PTCs. In node metastases, BRAF mutations were, in most but not all the cases, concordant with the dominant tumor. A discordant pattern of BRAF mutation was also found in about 50% of the cases in which multiple foci of different histopathological variants were present. CONCLUSIONS: The heterogeneous distribution of BRAF mutations suggests that discrete tumor foci in multifocal PTC may occur as independent tumors. This information has to be considered in the design of targeted therapeutic approaches with BRAF pathway inhibitors.