Jaroslaw Jendrzejewski

The polymorphism rs944289 predisposes to papillary thyroid carcinoma through a large intergenic noncoding RNA gene of tumor suppressor type

A genome-wide association study of papillary thyroid carcinoma (PTC) pinpointed two independent SNPs (rs944289 and rs965513) located in regions containing no annotated genes (14q13.3 and 9q22.33, respectively). Here, we describe a unique, long, intergenic, noncoding RNA gene (lincRNA) named Papillary Thyroid Carcinoma Susceptibility Candidate 3 (PTCSC3) located 3.2 kb downstream of rs944289 at 14q.13.3 and the expression of which is strictly thyroid specific. By quantitative PCR, PTCSC3 expression was strongly down-regulated (P = 2.84 × 10−14) in thyroid tumor tissue of 46 PTC patients and the risk allele (T) was associated with the strongest suppression (genotype [TT] (n = 21) vs. [CT] (n = 19), P = 0.004). In adjacent unaffected thyroid tissue, the genotype [TT] was associated with up-regulation of PTCSC3 ([TT] (n = 21) vs. [CT] (n = 19), P = 0.034). The SNP rs944289 was located in a binding site for the CCAAT/enhancer binding proteins (C/EBP) α and β. The risk allele destroyed the binding site in silico. Both C/EBPα and C/EBPβ activated the PTCSC3 promoter in reporter assays (P = 0.0009 and P = 0.0014, respectively) and the risk allele reduced the activation compared with the nonrisk allele (C) (P = 0.026 and P = 0.048, respectively). Restoration of PTCSC3 expression in PTC cell line cells (TPC-1 and BCPAP) inhibited cell growth (P = 0.002 and P = 0.019, respectively) and affected the expression of genes involved in DNA replication, recombination and repair, cellular movement, tumor morphology, and cell death. Our data suggest that SNP rs944289 predisposes to PTC through a previously uncharacterized, long intergenic noncoding RNA gene (PTCSC3) that has the characteristics of a tumor suppressor.

PTCSC3 Is Involved in Papillary Thyroid Carcinoma Development by Modulating S100A4 Gene Expression

CONTEXT We previously showed that a long noncoding RNA gene, PTCSC3, located close to the variant rs944289 that predisposes to papillary thyroid carcinoma (PTC) might target the S100A4 gene. OBJECTIVE The aim was to investigate the impact of PTCSC3 on S100A4 expression and its role in cancer development. DESIGN S100A4 abundance was analyzed by quantitative PCR (qPCR) in unaffected and tumor tissue from n = 73 PTC patients. The expression of PTCSC3 and S100A4 was studied in BCPAP and TPC-1 cell lines with forced expression of PTCSC3 by qPCR. Expression of S100A4 target genes (VEGF and MMP-9) was studied in the BCPAP cell line with forced expression of PTCSC3 by qPCR, reverse transcriptase PCR, and Western blot. The impact of PTCSC3 on BCPAP motility and invasiveness was analyzed by the Transwell and Matrigel assays, respectively. SETTING This was a laboratory-based study using cells from clinical samples and thyroid cancer cell lines. MAIN OUTCOME AND MEASURE We aimed to find evidence for a link between the expression of PTCSC3 and thyroid carcinogenesis. RESULTS Expression data from PTC cell lines pinpointed S100A4 as the most significantly downregulated gene in the presence of PTCSC3. S100A4 was upregulated in tumor tissue (P = 9.33 × 10(-7)) while PTCSC3 was strongly downregulated (P = 2.2 × 10(-16)). S100A4 transcription was moderately correlated with PTCSC3 expression in unaffected thyroid tissue (r = 0.429, P = .0001), and strongly in unaffected tissue of patients with the risk allele of rs944289 (r = 0.685, P = 7.88 × 10(-5)). S100A4, VEGF, and MMP-9 were suppressed in the presence of PTCSC3 (P = .0051, P = .0090, and P =.0037, respectively). PTC cells expressing PTCSC3 showed reduction in motility and invasiveness (P = 4.52 × 10(-5) and P = 1.0 × 10(-4), respectively). CONCLUSIONS PTCSC3 downregulates S100A4, leading to a reduction in cell motility and invasiveness. We propose that PTCSC3 impacts PTC predisposition and carcinogenesis through the S100A4 pathway.

Ultra-rare mutation in long-range enhancer predisposes to thyroid carcinoma with high penetrance.

Thyroid cancer shows high heritability but causative genes remain largely unknown. According to a common hypothesis the genetic predisposition to thyroid cancer is highly heterogeneous; being in part due to many different rare alleles. Here we used linkage analysis and targeted deep sequencing to detect a novel single-nucleotide mutation in chromosome 4q32 (4q32A>C) in a large pedigree displaying non-medullary thyroid carcinoma (NMTC). This mutation is generally ultra-rare; it was not found in 38 NMTC families, in 2676 sporadic NMTC cases or 2470 controls. The mutation is located in a long-range enhancer element whose ability to bind the transcription factors POU2F and YY1 is significantly impaired, with decreased activity in the presence of the C- allele compared with the wild type A-allele. An enhancer RNA (eRNA) is transcribed in thyroid tissue from this region and is greatly downregulated in NMTC tumors. We suggest that this is an example of an ultra-rare mutation predisposing to thyroid cancer with high penetrance.

Telomere Length and Telomerase Reverse Transcriptase Gene Copy Number in Patients with Papillary Thyroid Carcinoma

CONTEXT The family risk ratio for papillary thyroid carcinoma (PTC) is among the highest of all cancers. Collectively, familial cases (fPTC) and sporadic cases (sPTC) are not known to show molecular differences. However, one study reported that telomeres were markedly shorter and the telomerase reverse transcriptase (TERT) gene was amplified and up-regulated in germline DNA from patients with fPTC compared with sPTC. OBJECTIVE The aim of this study was to evaluate telomere length and TERT gene amplification and expression in blood samples of fPTC and sPTC patients in a genetically distinct population from the previous study. DESIGN In 42 fPTC and 65 sPTC patients, quantitative real-time PCR was employed to measure the relative telomere length (RTL) and TERT gene copy number and RNA level. To validate the results using alternative methods, we further studied a subset of the original cohort consisting of randomly chosen fPTC (n = 10) and sPTC (n = 14) patients and controls (n = 21) by assessing both telomere length by flow fluorescent in situ hybridization and TERT gene expression by quantitative real-time PCR. RESULTS RTL and TERT gene copy number did not differ between fPTC and sPTC (P = 0.957 and P = 0.998, respectively). The mean RTL and TERT gene expression were not significantly different among the groups of the validation series (P = 0.169 and P = 0.718, respectively). CONCLUSION Our data show no difference between familial and sporadic PTC with respect to telomere length, TERT copy number, or expression in our cohort. Further investigations in additional cohorts of patients are desirable.

Pheochromocytoma during pregnancy – successful surgical treatment

322 showed a mean BP of 199/125 mm Hg, mean daytime BP of 202/130 mm Hg, mean noctur‐ nal BP of 194/115 mm Hg, and a maximum BP of 230/142 mm Hg. Diagnostic workup for secondary hyperten‐ sion included the measurement of plasma renin activity and 24 ‐hour urine collection. The urine collection revealed a markedly elevated excretion of normetanephrine (5760 μg/24 h; reference range, 162–527 μg/24 h) and 3 ‐methoxytyramine (1817 μg/24 h; reference range, 103–434 μg/24 h). Abdominal ultrasound showed a pathological mass in the left adrenal gland. Magnetic reso‐ nance imaging confirmed the presence of a large heterogeneous adrenal tumor of 80 × 62 × 68 mm in size (FIGURE 1A and 1B). Considering the clinical picture and radio‐ logical and laboratory test results, pheochro‐ mocytoma (PCC) of the left adrenal gland was A 30 ‐year ‐old woman at 12 weeks of gestation presented to the hospital with high blood pres‐ sure (BP) (maximum, 225/136 mm Hg) despite antihypertensive treatment with metoprolol, ni‐ trendipine, and methyldopa. She had a 4 ‐year history of arterial hypertension. Abdominal ul‐ trasound and dynamic renal scintigraphy per‐ formed 2 years earlier revealed no abnormali‐ ties. Her family history was negative for hyper‐ tension and cardiovascular diseases. On admis‐ sion, she presented with anxiety, palpitations, and recurring headaches. Physical examination revealed a BP of 200/140 mm Hg and a heart rate (HR) of 140 bpm, as well as systolic heart mur‐ mur. Laboratory tests at baseline showed pro‐ teinuria, gestational diabetes, and normal thyroid hormone levels. Electrocardiography revealed si‐ nus tachycardia (127 bpm). There were no abnor‐ malities on echocardiography. Holter monitoring CLINICAL IMAGE

Fine mapping of 14q13 reveals novel variants associated with different histological subtypes of papillary thyroid carcinoma: PTC associated 14q13 variants

The first two genome wide association studies (GWAS) of papillary thyroid carcinoma (PTC) detected five variants associated with PTC. Two of them (rs944289 and rs116909374) are located at 14q13 making that locus an important target of research into the genetic predisposition to PTC. We aimed at uncovering other variants at 14q13 associated with PTC independently from the GWAS variants. We performed next generation sequencing of the 14q13 region and analyzed the allele frequencies of single nucleotide polymorphisms (SNPs) in n = 90 PTC cases vs. n = 379 EUR controls from the 1,000 Genome Project. The variants associated with PTC were validated in an Ohio cohort of n = 1,216 PTC cases and n = 1,416 controls. Next, we analyzed the association between SNPs and expression of nearby genes and clinical parameters. We showed that rs368187 was associated with PTC (OR = 1.31, p = 2.20 × 10−6). Rs1632250, Rs1863347 and rs1755787 showed association with classical PTC (cPTC) (n = 891; OR = 1.24, 2.22 × 10−3, OR = 1.31, p = 2.15 × 10−4 and OR = 1.24, p = 2.06 × 10−3, respectively) while variant rs28397092 showed association with follicular variant (n = 243; OR = 1.51, p = 1.36 × 10−3). Rs1863347 was associated with suppression of PTCSC3 in unaffected thyroid tissue (p = 0.026). Rs1632250, rs1863347 and rs1755787 showed association with multifocality (OR = 1.85, p = 0.001, OR = 1.98, p = 0.001 and OR = 1.76, p = 0.003 respectively) and N stage (OR = 1.79, p = 0.014, OR = 1.73, p = 0.023 and OR = 1.81, p = 0.013, respectively) in microPTC (n = 328) while rs368187 was associated with M stage (OR = 0.56, p = 0.034) in cPTC. Our results disclose multiple variants associated with PTC and clinical features in the 14q13 superlocus. We suggest that translational genotype/phenotype studies should take into account not only somatic mutations but also germline variants.

Correction: Ultra-Rare Mutation in Long-Range Enhancer Predisposes to Thyroid Carcinoma with High Penetrance

An additional grant from the National Cancer Institute was incorrectly omitted from the Funding Statement. The number of the grant is: R01 CA151979.

Thyroid Hormone Receptor β (THRB) Is a Major Target Gene for Micro-RNAs Deregulated in Papillary Thyroid Carcinoma (PTC)

This article appears in The Journal of Clinical Endocrinology & Metabolism, published December 15, 2010, 10.1210/jc.2010-1594