Recent advances in circulating nucleic acids in oncology.

Circulating cell-free DNA (cfDNA) is one of the fastest growing and most exciting areas in oncology in recent years. Its potential clinical uses cover now each phase of cancer patient management care (predictive information, detection of the minimal residual disease, early detection of resistance, treatment monitoring, recurrence surveillance, and cancer early detection/screening). This review relates the recent advances in the application of circulating DNA or RNA in oncology building on unpublished or initial findings/work presented at the 10th international symposium on circulating nucleic acids in plasma and serum held in Montpellier from the 20th to the 22nd of September 2017. This year, presenters revealed their latest data and crucial observations notably in relation to (i) the circulating cell-free (cfDNA) structure and implications regarding their optimal detection; (ii) their role in the metastatic or immunological processes; (iii) evaluation of miRNA panels for cancer patient follow up; (iv) the detection of the minimal residual disease; (v) the evaluation of a screening tests for cancer using cfDNA analysis; and (vi) elements of preanalytical guidelines. This work reviews the recent progresses in the field brought to light in the meeting, as well as in the most important reports from the literature, past and present. It proposes a broader picture of the basic research and its potential, and of the implementation and current challenges in the use of circulating nucleic acids in oncology.

Circulating cell-free cancer-testis MAGE-A RNA, BORIS RNA, let-7b and miR-202 in the blood of patients with breast cancer and benign breast diseases.

BACKGROUND: MAGE-A (melanoma-associated antigen-A) are promising targets for specific immunotherapy and their expression may be induced by the epigenetic factor BORIS. METHODS: To determine their relevance for breast cancer, we quantified the levels of MAGE-A1, -A2, -A3, -A12 and BORIS mRNA, as well as microRNAs let-7b and miR-202 in pre- and postoperative serum of 102 and 34 breast cancer patients, respectively, and in serum of 26 patients with benign breast diseases and 37 healthy women by real-time PCR. The mean follow-up time of the cancer patients was 6.2 years. RESULTS: The serum levels of MAGE-A and BORIS mRNA, as well as let-7b were significantly higher in patients with invasive carcinomas than in patients with benign breast diseases or healthy women (P<0.001), whereas the levels of miR-202 were elevated in both patient cohorts (P<0.001). In uni- and multivariate analyses, high levels of miR-202 significantly correlated with poor overall survival (P=0.0001). Transfection of breast cancer cells with synthetic microRNAs and their inhibitors showed that let-7b and miR-202 did not affect the protein expression of MAGE-A1. CONCLUSIONS: Based on their cancer-specific increase in breast cancer patients, circulating MAGE-A and BORIS mRNAs may be further explored for early detection of breast cancer and monitoring of MAGE-directed immunotherapies. Moreover, serum miR-202 is associated with prognosis.

[New aspects in sonography of the liver]. / Neue Horizonte bei der Lebersonographie.

Ultrasound-based technologies are competing more and more success-fully against computerized tomography and magnetic resonance based imaging procedures. This thanks to technological improvements as well as accumulating evidence, stemming from properly conducted controlled clinical studies. The reduced cost and radiation safety issues are additional arguments in its favor. This article reviews new developments in ultrasound medicine as applied to the liver. Emphasis lies on new data related to contrast-ultrasound (Sulphur Hexafluoride SonoVue®) which allows a dynamic analysis of liver perfusion and hence improved characterization of focal liver lesions, such as metastases of extrahepatic tumors, regenerative nodules in patients with liver cirrhosis, focal nodular hyperplasia, hepatocellular carcinoma, liver hemangioma, liver adenoma and or focal hypo-respectively hypersteatosis. This article also deals with important new techniques, which allow assessment of liver stiffness such as transient elastography (Fibroscan), ARFI (Acoustic Radiation Force Impulse) or real-time-tissue elastography. These new techniques will help us to assess and quantify the levels of liver steatosis with more precision and permit accurate follow-up measurements.

[Diagnosis and management of suspected nephrolithiasis in a primary care setting]. / Die Nierenkolik in der allgemein-internistischen Praxis.

Based on the prevalence of asymptomatic kidney stones (5% in our general ward, in accordance with the literature) the value of abdominal ultrasonography in the clinical assessment of a suspected kidney-colic is discussed. The eminent importance of the stone-analysis is emphasized. In addition, the most common causes of kidney stone formation (low urine output, mechanical urinary obstruction in the renal pelvis, hypercalciuria, hyperoxaluria, insufficient urinary citric acid excretion, hyperuricosuria) are highlighted. The cardinal symptom of the urolithiasis is the presence of micro/macrohematuria (which is often absent - according to citations - in 20-80%!). Moreover, the differential diagnosis of acute flank pain, as neoplastic- or infectious diseases, reno-vascular and extrarenal causes (retro-peritoneal and mesenteric vascular processes and rupture of abdominal aneurysms), gynecological problems (e.g. rotation/rupture of ovarian cysts, ectopic pregnancy), appendicitis, diverticulitis, and splenic abscess/infarction, as well as hepato-pancreaticobiliary causes are discussed. Moreover, metabolic syndromes, e.g. the intermittant porphyria or infectious diseases (e.g, Fitz-Hugh-Curtis syndrome) and other rare pathologies (such as the «Mediterranean fever¼) may be at the origin of acute flank pains. A particular attention is given to possible diagnostic procedures in a primary care setting: in addition to medical history, clinical status and specific laboratory findings the value of diagnostic ultrasound, with special reference to the color-Doppler application, as the «twinkling artefact¼ from kidney stones and the «urinary-jet phenomenon¼ for the assessment of urinary outflow obstruction, is emphasized. In this context we point out that a lack of dilatation of the kidney pelvis never excludes a kidney-colic, on the other hand, a dilatation of the kidney pelvis does not necessarily mean congestion! The conservative treatment strategies (avoidance of excessive drinking - an obstructed kidney protects itself - NSAID in combination with Tamsulosin, especially in case of prevesical urolithiasis) are discussed. The critical stone size (≤5 mm) and the absence of «red flags¼ (especially obstructive and inflammatory signs) allow a non-specialist medical outpatient treatment of acute nephro-and ureterolithiasis. The possible complications of the urolithiasis, especially the urosepsis and the (iatrogenic) fornix rupture are highlighted, as well as the formation of a renal abscess or hydronephrosis. A short look is given to the metaphylaxis of the urolithiasis and its «recurrence rate¼.

[Ultrasound in the musculoskeletal system - possibilities and limits]. / Der Ultraschall am Bewegungsapparat - Möglichkeiten und Grenzen.

Since the early 1990's, ultrasound through the development of high-resolution probes in the musculoskeletal system has become more and more important. The development of ultrasound technology has gone hand in hand with the development of know-how in different clinical fields, so that diagnostic ultrasound has now become an indispensable diagnostic tool in orthopedics/traumatology, rheumatology and neurology. The following article will arouse curiosity and interest in musculoskeletal ultrasound.

[«Knobology¼ (important device settings)]. / «Knopfologie¼ (Wichtige Geräteeinstellungen).

The correct setting of the ultrasound equipment is a prerequisite for a high quality of the ultrasound examination. Both the B-mode ultrasonography, as well as the Doppler technique offer interesting alternative settings that are often inadequately used. This brief summary of basic settings should help the practicing clinicians in everyday life and motivate them to deal with the «knobs¼ of the ultrasound device. Good practical help will provided by the «Sonocave¼ an event regularly organized at the annual ultrasonic training Congress in Davos, where recognized experts will answer such «Knobology¼-questions and demonstrate the appropriate settings also practically.

[Image analysis in the differential diagnosis of renal parenchyma lesions]. / Der Beitrag der Bildanalyse für die Differenzialdiagnose der fokalen Nierenparenchymveränderungen.

PURPOSE: Visual analysis of echo intensity is of importance for the differential diagnosis of focal renal lesions. Quantification of the echo intensity and of other parameters might help with differential diagnosis. MATERIALS AND METHODS: In 145 patients with focal renal lesions, digitized images were evaluated (40 angiomyolipomas [group A], 70 renal cell carcinomas [group B], 20 pseudo-tumors [group C] and 15 other focal lesions in group D). With Photoshop®, the average grayscale values of the lesion (defined as echo intensity focal, EF) and its variance s2 (as expression of the inhomogeneity focal, IF) were measured. These measurements were compared to the renal cortex (echo intensity renal cortex=ER, inhomogeneity renal renal cortex=I R). Other calculated parameters: Echo intensity quotient, echo intensity index, inhomogeneity quotient and inhomogeneity index. RESULTS: Angiomyolipomas had a higher echo intensity quotient EQ and echo intensity index EI than renal cell carcinomas, pseudo-tumors and other lesions (p<0.001). Pseudo-tumors had a lower inhomogeneity quotient than angiomyolipomas (p<0.001), renal cell carcinomas (p<0.05). Echo intensity quotient EQ≥2.0 and echo intensity index EI≥0.5 were typical for angiomyolipomas with a sensitivity of 96.4 % and a specificity of 97.3 % for tumors<3 cm. CONCLUSION: Quantitative echo intensity measurements enhance the differential diagnosis of focal renal lesions. The differentiation of typical angiomyolipomas to other lesions could be improved.

[The quantitative measurement of the echogenicity of the renal parenchyma]. / Die quantitative Messung der Echogenität des Nierenparenchyms.

The qualitative assessment of the echogenicity of the renal cortex and the medulla is one of the most important diagnostic criteria in the assessment of diffuse renal parenchymal disease. It is of interest to complete this assessment by quantitative data. The echogenicity of the cortex and medulla was quantitatively analysed in digitized images. The coefficient of variation of repeated measurements was 0.83% and the coefficient of variation made by two different individuals was 2.03%. The influence of furosemide on the echogenicity of the renal parenchyma was measured in 4 healthy adults. The echogenicity of the renal cortex is after 3 and 6 min significantly increased, while the echogenicity of the medulla remained unchanged. Our study shows that a precise measurement of echogenicity in renal parenchyma is possible. Under the influence of furosemide there will be a significant increase in the echogenicity of the renal cortex.

[Ultrasound for renal colic]. / Die Sonographie bei Nierenkolik.

Nowadays the Spiral-CT-scan is stressed as the diagnostic "gold-standard" in the management of kidney-colics. Our findings suggest a very good sensitivity (98%) and specificity of diagnostic ultrasound (items:Hydrourether, RI > 0.05) in combination with hematuria in an ambulatory setting. Our data are referred to 70 ambulatory patients examined with the hypothetical diagnosis of kidney-colic in a general medicine surgery between 1996-2003. Diagnostic ultrasound is a reliable tool in the management of kidney-colics in ambulatory medicine. Its cost-saving character and the lack of X-rays-exposure should be taken into account in choosing the appropriate method in judging abdominal pains possibly related to kidney-colics.

[Refreshing in abdominal ultrasonography--a summary from SGUM Kongress Davos in 2003]. / Stellenwert der abdominellen Sonographie Refresherkurs im Rahmen des SGUM-Kongresses.

Ultrasonography is a modern noninvasive imaging technique with a high resolution, and high evidence in diagnostic questions, high acceptance in patients, low costs and no side effects. There are pure questions in whitch the diagnostic ultrasound is not able to give a satisfactory answer. Important is basically a well trained examinator with long experience and an exact examination and documentation.

Gene structure and characterization of the murine homologue of the B cell-specific transcriptional coactivator OBF-1.

The B cell-specific activity of immunoglobulin (Ig) gene promoters is to a large extent mediated by the conserved octamer motif ATTTGCAT. This requires the DNA binding octamer factors Oct-1 and/or Oct-2, as well as an additional B cell-restricted non-DNA binding cofactor. We recently cloned such a coactivator specific for Oct-1 or Oct-2 from human B cells and called it OBF-1. Here we report the isolation and characterization of the murine homologue. Full-length cDNA clones as well as genomic clones were isolated and the gene structure was determined. The deduced protein sequence shows that the mouse protein has an identical length, is likewise proline rich and shows 89% overall identity to the human protein. The OBF-1 gene is expressed in a very highly B cell-specific manner and is transcribed in cells representative of all stages of B cell differentiation, including the earliest ones. We show that OBF-1 interacts in the absence of DNA with the POU domain of Oct-1 or Oct-2 and also with the general transcription factors TBP and TFIIB. Furthermore, we demonstrate that although OBF-1 efficiently activates promoter octamer sites, it does not activate enhancer octamer sites.

Involvement of the Ets family factor PU.1 in the activation of immunoglobulin promoters.

The B cell-specific expression of immunoglobulin (Ig) genes is controlled by the concerted action of variable (V) region promoters and intronic or 3' enhancers, all of which are active in a lymphoid-specific manner. A crucial highly conserved element of the V region promoters is the octamer site -ATTTGCAT-, which can be bound by ubiquitous (Oct-1) as well as B cell-specific (Oct-2) factors. Another less conserved element found in many Ig promoters is pyrimidine-rich and has been shown to be functionally important, in particular for those Ig promoters that have only an imperfect octamer site. In this study we have analyzed the factors binding specifically to the pyrimidine-rich motif of the V kappa 19 promoter, a light chain gene promoter with an imperfect octamer site. Using nuclear extracts prepared from B cells, we detected two sets of specific complexes in electrophoretic mobility shift experiments. One complex appears to be ubiquitous but enriched in lymphoid cells and represents the binding of a potentially novel factor with an apparent molecular mass of approximately 50 kDa. The other complex was found only with extracts from pre-B or B cells as well as from a macrophage cell line and appears to be caused by the binding of PU.1, a factor of the Ets family. We show that on this Ig promoter Oct factors (Oct-1 or Oct-2) and PU.1 can bind concomitantly but without synergism. By transfection experiments in non-B cells we demonstrate that PU.1 is indeed able to activate this promoter in concert with Oct-2. Furthermore, we show that PU.1 can bind with varying affinities to the pyrimidine-rich elements of several other Ig promoters. These data suggest a more general role for PU.1 or other members of the Ets family in the activation of Ig promoters.

Octamer factors exert a dual effect on the IL-2 and IL-4 promoters.

The promoters of IL-2 and IL-4 genes contain multiple binding sites for octamer factors. In peripheral T lymphocytes and several T cell lines, both the ubiquitous Oct factor Oct-1 and the lymphocyte-specific factor Oct-2 are expressed and bind to the IL-2 and IL-4 promoters. Prominent octamer binding sites of IL-2 and IL-4 promoters are their upstream promoter sites (UPS) which share 14 identical nucleotides. Multiple copies of the IL-2 and IL-4 UPS act as inducible enhancers in T cells, and their induction is inhibited by the immunosuppressant cyclosporin A (CsA). Closely linked to the octamer site, the IL-2 UPS contains a non-canonical AP-1 binding (TRE) site, and mutation in either site to a non-functional factor binding site impairs the induction of the IL-2 promoter. The binding of AP-1 and octamer factors to the IL-2 UPS DNA overlaps, and the tight association and functional cooperation of octamer with AP-1 factors is of crucial importance for the inducible IL-2 UPS activity. Introduction of five or ten spacer nucleotides between both IL-2 UPS sites results in a drastic reduction of inducible UPS activity, both in the loss of suppression by CsA and stimulation by the Ca(2+)-dependent phosphatase calcineurin. Within the IL-4 UPS the Oct and TRE-like motifs are separated by a binding site of nuclear factor of activated T cells (NF-AT). This site shares nine out of ten bp with an IL-2 NF-AT site. The strong binding of NF-ATp to the IL-4 UPS site suppresses the simultaneous binding of Oct factors to the IL-4 UPS. Because the two other Oct binding sites of IL-4 promoter show a similar sequence configuration, the binding of NF-AT seems to prevent the simultaneous binding of Oct factors to the IL-4 promoter. By contrast, both classes of factors bind simultaneously to the IL-2 promoter, and their tight association with AP-1 enhances the IL-2 promoter activity.

Interleukin 1 and tumor necrosis factor stimulate cGMP formation in rat renal mesangial cells.

Treatment of mesangial cells with recombinant human interleukin 1 beta (IL-1 beta) or recombinant human tumor necrosis factor alpha (TNF alpha) dose-dependently increased cGMP formation. Both IL-1 beta and TNF alpha-stimulated formation of cGMP occurred after a initial lag period of 4 to 8 hours. Treatment of cells with actinomycin D, cycloheximide or dexamethason completely abolished cytokine-induced cGMP formation. Furthermore, the guanylate cyclase inhibitor Methylene blue completely blocked IL-1 beta- and TNF alpha-stimulated cGMP generation. NG-mono-methyl-L-arginine attenuated IL-1 beta- and TNF alpha-induced cGMP production, an effect that was reversed by L-arginine.

[1,200 mg rifampicin(Rimactan) as an oral one-dose treatment of uncomplicated gonorrhea in women]. / 1200 mg Rifampicin (Rimactan) als Einzeldosisbehandlung der unkomplizierten Gonorrhoe der Frau.

40 women were treated for uncomplicated genital gonorrhoea, verified by culture, with a single oral dose of 1,200 mg rifampicin (Rimactan). The ages ranged between 15 and 51 years. In 33 patients that could be evaluated, the gonorrhoea was cured. In 8 of 18 patients the preexistent vaginal discharge improved. 4 women experienced six different side effects which were probably related to rifampicin. The 1,200-mg rifampicin oral one-dose treatment is a reliable, efficient, and well-tolerated therapy of uncomplicated gonorrhoea in women.

Skin reactivity, basophil degranulation and IgE levels in ageing.

Skin tests with histamine, the histamine-liberator, codeine, and various allergens as well as blood basophil degranulation by anti-IgE or anti-IgG4 and total serum IgE levels have been studied in two female populations of different ages (average 23.1 and 73.9 years). All thirty-one patients selected for this study were clinically non-allergic. We observed a trend towards reduced skin reactions to histamine and codeine in the higher age-group; on the other hand, we could not find any decrease of basophil degranulation as a sign of basophil impairment with age. Likewise no difference in total serum IgE levels have been noticed. No correlation between skin tests and basophil degranulation was observed; yet patients with isolated, positive skin tests to house dust and/or Candida albicans showed a statistically significant reduced blood basophil degranulation by anti-IgE or anti-IgG4.

[Plasma exchange in pemphigus vulgaris]. / Plasmaaustausch bei Pemphigus vulgaris.

An attempt to treat a patient with pemphigus vulgaris by large volume plasma exchange with intravenous application of high doses of immunoglobulin at the end of each exchange procedure is reported. The therapeutic results--temporary clinical improvement parallel to a significant reduction in the level of circulating antiepithelial autoantibodies--are discussed in comparison with published case reports on plasma exchange in pemphigus vulgaris and bullous pemphigoid.