Attila Patócs

A prospective, multicentre study to investigate the efficacy, safety and tolerability of octreotide LAR® (long-acting repeatable octreotide) in the primary therapy of patients with acromegaly

Objective  To evaluate the efficacy, safety and tolerability of octreotide LAR® (long‐acting repeatable octreotide) in the primary therapy of acromegaly.

Germline Mutations and Variants in the Succinate Dehydrogenase Genes in Cowden and Cowden-like Syndromes

Individuals with PTEN mutations have Cowden syndrome (CS), associated with breast, thyroid, and endometrial neoplasias. Many more patients with features of CS, not meeting diagnostic criteria (termed CS-like), are evaluated by clinicians for CS-related cancer risk. Germline mutations in succinate dehydrogenase subunits SDHB-D cause pheochromocytoma-paraganglioma syndrome. One to five percent of SDHB/SDHD mutation carriers have renal cell or papillary thyroid carcinomas, which are also CS-related features. SDHB-D may be candidate susceptibility genes for some PTEN mutation-negative individuals with CS-like cancers. To address this hypothesis, germline SDHB-D mutation analysis in 375 PTEN mutation-negative CS/CS-like individuals was performed, followed by functional analysis of identified SDH mutations/variants. Of 375 PTEN mutation-negative CS/CS-like individuals, 74 (20%) had increased manganese superoxide dismutase (MnSOD) expression, a manifestation of mitochondrial dysfunction. Among these, 10 (13.5%) had germline mutations/variants in SDHB (n = 3) or SDHD (7), not found in 700 controls (p < 0.001). Compared to PTEN mutation-positive CS/CS-like individuals, those with SDH mutations/variants were enriched for carcinomas of the female breast (6/9 SDH versus 30/107 PTEN, p < 0.001), thyroid (5/10 versus 15/106, p < 0.001), and kidney (2/10 versus 4/230, p = 0.026). In the absence of PTEN alteration, CS/CS-like-related SDH mutations/variants show increased phosphorylation of AKT and/or MAPK, downstream manifestations of PTEN dysfunction. Germline SDH mutations/variants occur in a subset of PTEN mutation-negative CS/CS-like individuals and are associated with increased frequencies of breast, thyroid, and renal cancers beyond those conferred by germline PTEN mutations. SDH testing should be considered for germline PTEN mutation-negative CS/CS-like individuals, especially in the setting of breast, thyroid, and/or renal cancers.

Integrative molecular bioinformatics study of human adrenocortical tumors: microRNA, tissue-specific target prediction, and pathway analysis

MicroRNAs (miRs) are involved in the pathogenesis of several neoplasms; however, there are no data on their expression patterns and possible roles in adrenocortical tumors. Our objective was to study adrenocortical tumors by an integrative bioinformatics analysis involving miR and transcriptomics profiling, pathway analysis, and a novel, tissue-specific miR target prediction approach. Thirty-six tissue samples including normal adrenocortical tissues, benign adenomas, and adrenocortical carcinomas (ACC) were studied by simultaneous miR and mRNA profiling. A novel data-processing software was used to identify all predicted miR-mRNA interactions retrieved from PicTar, TargetScan, and miRBase. Tissue-specific target prediction was achieved by filtering out mRNAs with undetectable expression and searching for mRNA targets with inverse expression alterations as their regulatory miRs. Target sets and significant microarray data were subjected to Ingenuity Pathway Analysis. Six miRs with significantly different expression were found. miR-184 and miR-503 showed significantly higher, whereas miR-511 and miR-214 showed significantly lower expression in ACCs than in other groups. Expression of miR-210 was significantly lower in cortisol-secreting adenomas than in ACCs. By calculating the difference between dCT(miR-511) and dCT(miR-503) (delta cycle threshold), ACCs could be distinguished from benign adenomas with high sensitivity and specificity. Pathway analysis revealed the possible involvement of G2/M checkpoint damage in ACC pathogenesis. To our knowledge, this is the first report describing miR expression patterns and pathway analysis in sporadic adrenocortical tumors. miR biomarkers may be helpful for the diagnosis of adrenocortical malignancy. This tissue-specific target prediction approach may be used in other tumors too.

Crosstalk between TGF-β signaling and the microRNA machinery.

The activin/transforming growth factor-β (TGF-β) pathway plays an important role in tumorigenesis either by its tumor suppressor or tumor promoting effect. Loss of members of the TGF-β signaling by somatic mutations or epigenetic events, such as DNA methylation or regulation by microRNA (miRNA), may affect the signaling process. Most members of the TGF-β pathway are known to be targeted by one or more miRNAs. In addition, the biogenesis of miRNAs is also regulated by TGF-β both directly and through SMADs. Based on these interactions, it appears that autoregulatory feedback loops between TGF-β and miRNAs influence the fate of tumor cells. Our aim is to review the crosstalk between TGF-β signaling and the miRNA machinery to highlight potential novel therapeutic targets.

Menin and its interacting proteins: elucidation of menin function

The multiple endocrine neoplasia type 1 (MEN1) gene is a tumor suppressor gene encoding a 610 amino acid nuclear protein, menin. Although mutations of the MEN1 gene are responsible for MEN 1 syndrome, the intracellular functions of menin have not been fully elucidated. Recent data suggest that interactions between menin and menin-interacting proteins have a role in physiological regulation of cell growth, control of the cell cycle and genome stability, and are potentially important in bone development and multipotent mesenchymal stem cell differentiation. Loss of these interactions might also contribute to the development of MEN 1 syndrome.

Risk Profiles and Penetrance Estimations in Multiple Endocrine Neoplasia Type 2A Caused by Germline RET Mutations Located in Exon 10

Multiple endocrine neoplasia type 2 is characterized by germline mutations in RET. For exon 10, comprehensive molecular and corresponding phenotypic data are scarce. The International RET Exon 10 Consortium, comprising 27 centers from 15 countries, analyzed patients with RET exon 10 mutations for clinical‐risk profiles. Presentation, age‐dependent penetrance, and stage at presentation of medullary thyroid carcinoma (MTC), pheochromocytoma, and hyperparathyroidism were studied. A total of 340 subjects from 103 families, age 4–86, were registered. There were 21 distinct single nucleotide germline mutations located in codons 609 (45 subjects), 611 (50), 618 (94), and 620 (151). MTC was present in 263 registrants, pheochromocytoma in 54, and hyperparathyroidism in 8 subjects. Of the patients with MTC, 53% were detected when asymptomatic, and among those with pheochromocytoma, 54%. Penetrance for MTC was 4% by age 10, 25% by 25, and 80% by 50. Codon‐associated penetrance by age 50 ranged from 60% (codon 611) to 86% (620). More advanced stage and increasing risk of metastases correlated with mutation in codon position (609→620) near the juxtamembrane domain. Our data provide rigorous bases for timing of premorbid diagnosis and personalized treatment/prophylactic procedure decisions depending on specific RET exon 10 codons affected. Hum Mutat 31:1–8, 2010.

Total-Genome Analysis of BRCA1/2-Related Invasive Carcinomas of the Breast Identifies Tumor Stroma as Potential Landscaper for Neoplastic Initiation

We have shown that the tumor microenvironment of sporadic breast cancer is diverse in genetic alterations and contributes to the cancer phenotype. The dynamic morphology of the mammary gland might be of special interest in hereditary breast/ovarian cancer syndrome (HBOC). We hypothesized that hotspots of loss of heterozygosity or allelic imbalance (LOH/AI) within the tumor stroma of BRCA1/2-related breast cancers provide an impaired mammary stroma that could facilitate later malignant transformation of the breast epithelium. We conducted a total genome LOH/AI scan of DNA derived from the epithelium and stroma of 51 BRCA1/2-related breast cancers, using 372 microsatellite markers. We compared these data with those from a set of 134 sporadic breast cancers. HBOC-related breast cancers accumulated significantly more genetic alterations than did sporadic breast cancers. BRCA1/2-related breast cancer stroma showed LOH/AI at 59.7% of all loci analyzed, similar to the average frequency of LOH/AI observed in the epithelium (66.2%). This is remarkably different from sporadic breast cancers, for which the average epithelial LOH/AI frequency (36.7%) far exceeds the average stromal LOH/AI frequency (28.4%) (P=.03). We identified 11 hotspot loci of LOH/AI in the BRCA1/2 stroma, encompassing genes such as POLD1, which functions in DNA replication, and SDHB. In a subset of samples, enriched for BRCA1 cases, we found 45.0% overall LOH/AI in the stroma, which was significantly higher than the 41.8% LOH/AI observed in corresponding epithelium (P=.04). Together, our data indicate that, in HBOC-related breast cancers, the accumulation of genomic instability in the cancer stroma coincides with that in the neoplastic epithelium, and we postulate that such a genetically unstable stroma might facilitate a microenvironment that functions as a landscaper that promotes genomic instability in the epithelium and, subsequently, neoplastic transformation.

Head and neck paragangliomas: clinical and molecular genetic classification

Head and neck paragangliomas are tumors arising from specialized neural crest cells. Prominent locations are the carotid body along with the vagal, jugular, and tympanic glomus. Head and neck paragangliomas are slowly growing tumors, with some carotid body tumors being reported to exist for many years as a painless lateral mass on the neck. Symptoms depend on the specific locations. In contrast to paraganglial tumors of the adrenals, abdomen and thorax, head and neck paragangliomas seldom release catecholamines and are hence rarely vasoactive. Petrous bone, jugular, and tympanic head and neck paragangliomas may cause hearing loss. The internationally accepted clinical classifications for carotid body tumors are based on the Shamblin Class I–III stages, which correspond to postoperative permanent side effects. For petrous-bone paragangliomas in the head and neck, the Fisch classification is used. Regarding the molecular genetics, head and neck paragangliomas have been associated with nine susceptibility genes: NF1, RET, VHL, SDHA, SDHB, SDHC, SDHD, SDHAF2 (SDH5), and TMEM127. Hereditary HNPs are mostly caused by mutations of the SDHD gene, but SDHB and SDHC mutations are not uncommon in such patients. Head and neck paragangliomas are rarely associated with mutations of VHL, RET, or NF1. The research on SDHA, SDHAF2 and TMEM127 is ongoing. Multiple head and neck paragangliomas are common in patients with SDHD mutations, while malignant head and neck paraganglioma is mostly seen in patients with SDHB mutations. The treatment of choice is surgical resection. Good postoperative results can be expected in carotid body tumors of Shamblin Class I and II, whereas operations on other carotid body tumors and other head and neck paragangliomas frequently result in deficits of the cranial nerves adjacent to the tumors. Slow growth and the tendency of hereditary head and neck paragangliomas to be multifocal may justify less aggressive treatment strategies.

MicroRNA profile indicates downregulation of the TGFβ pathway in sporadic non-functioning pituitary adenomas

MicroRNAs (miRs) are small, 16–29 nucleotide long, non-coding RNA molecules which regulate the stability or translational efficiency of targeted mRNAs via RNA interference. MiRs participate in the control of cell proliferation, cell differentiation, signal transduction, cell death, and they play a role in carcinogenesis. The aims of our study were to analyse the expression profile of miRs in sporadic clinically non-functioning pituitary adenomas (NFPA) and in normal pituitary tissues, and to identify biological pathways altered in these pituitary tumors. MiR expression profiles of 12 pituitary tissue specimens (8 NFPA and 4 normal pituitary tissues) were determined using miR array based on quantitative real-time PCR with 678 different primers. Five overexpressed miRs and mRNA expression of Smads (Smad1-9), MEG and DLK1 genes were evaluated with individual Taqman assays in 10 NFPA and 10 normal pituitary tissues. Pathway analysis was performed by the DIANA-mirPath tool. Complex bioinformatical analysis by multiple algorithms and association studies between miRs, Smad3 and tumor size was performed. Of the 457 miRs expressed in both NFPA and normal tissues, 162 were significantly under- or overexpressed in NFPA compared to normal pituitary tissues Expression of Smad3, Smad6, Smad9, MEG and DLK1 was significantly lower in NFPA than in normal tissues. Pathway analysis together with in silico target prediction analysis indicated possible downregulation of the TGFβ signaling pathway in NFPA by a specific subset of miRs. Five miRs predicted to target Smad3 (miR-135a, miR-140-5p, miR-582-3p, miR-582-5p and miR-938) were overexpressed. Correlation was observed between the expression of seven overexpressed miRs and tumor size. Downregulation of the TGFβ signaling through Smad3 via miRs may have a possible role in the complex regulation of signaling pathways involved in the tumorigenesis process of NFPA.

Analysis of circulating microRNAs in adrenocortical tumors

Differential diagnosis of adrenocortical adenoma (ACA) and carcinoma is of pivotal clinical relevance, as the prognosis and clinical management of benign and malignant adrenocortical tumors (ACTs) is entirely different. Circulating microRNAs (miRNAs) are promising biomarker candidates of malignancy in several tumors; however, there are still numerous technical problems associated with their analysis. The objective of our study was to investigate circulating miRNAs in ACTs and to evaluate their potential applicability as biomarkers of malignancy. We have also addressed technical questions including the choice of profiling and reference gene used. A total of 25 preoperative plasma samples obtained from patients with ACAs and carcinomas were studied by microarray and quantitative real-time PCR. None of the three miRNAs (hsa-miR-192, hsa-mir-197 and hsa-miR-1281) found as differentially expressed in plasma samples in our microarray screening could be validated by quantitative real-time PCR. In contrast, of the selected eight miRNAs reported in the literature as differentially expressed in ACT tissues, five (hsa-miR-100, hsa-miR-181b, hsa-miR-184, hsa-miR-210 and hsa-miR-483-5p) showed a statistically significant overexpression in adrenocortical cancer vs adenoma when normalized on hsa-miR-16 as a reference gene. Receiver operator characteristic analysis of data revealed that the combination of dCThsa-miR-210 - dCThsa-miR-181b and dCThsa-miR-100/dCThsa-miR-181b showed the highest diagnostic accuracy (area under curve 0.87 and 0.85, respectively). In conclusion, we have found significant differences in expression of circulating miRNAs between ACAs and carcinomas, but their diagnostic accuracy is not yet high enough for clinical application. Further studies on larger cohorts of patients are needed to assess the diagnostic and prognostic potential application of circulating miRNA markers.