High risk of hypogonadism in young male cancer survivors.

OBJECTIVE: Cancer and its treatment in childhood and young adulthood can cause hypogonadism, leading to increased risk of long-term morbidity and mortality. The aim of this study was to evaluate the risk of presenting with biochemical signs of hypogonadism in testicular cancer survivors (TCS) and male childhood cancer survivors (CCS) in relation to the type of treatment given. DESIGN: Case-control study. PATIENTS: Ninety-two TCS, 125 CCS (mean age 40 and median age 34 years, respectively; mean follow-up time 9.2 and 24 years, respectively) and a corresponding number of age-matched controls. MEASUREMENTS: Fasting morning blood samples were analysed for total testosterone (TT), follicle-stimulating hormone (FSH) and luteinizing hormone (LH). The odds ratios (OR) for hypogonadism, defined as primary, secondary, compensated or ongoing androgen replacement, were calculated for TCS and CCS and for subgroups defined by diagnosis and treatment. RESULTS: Hypogonadism was found in 26% of CCS and 36% of TCS, respectively (OR: 2.1, P = .025 and OR = 2.3, P = .021). Among CCS, the OR was further increased in those given testicular irradiation (OR = 28, P = .004). Radiotherapy other than cranial or testicular irradiation plus chemotherapy, or cranial irradiation without chemotherapy, associated also with increased ORs (OR = 3.7, P = .013, and OR = 4.4, P = .038, respectively). Among TCS, those receiving >4 cycles of cisplatin-based chemotherapy had OR = 17, P = .015. CONCLUSIONS: Biochemical signs of testosterone deficiency are recognized as markers of decreased life expectancy. Thus, the risk of hypogonadism in TCS and CCS should be recognized and emphasizes the need of long-term follow-up for these men.

First Experimental Characterization of Microwave Emission from Cosmic Ray Air Showers.

We report the first direct measurement of the overall characteristics of microwave radio emission from extensive air showers. Using a trigger provided by the KASCADE-Grande air shower array, the signals of the microwave antennas of the Cosmic-Ray Observation via Microwave Emission experiment have been read out and searched for signatures of radio emission by high-energy air showers in the GHz frequency range. Microwave signals have been detected for more than 30 showers with energies above 3×10^{16} eV. The observations presented in this Letter are consistent with a mainly forward-directed and polarized emission process in the GHz frequency range. The measurements show that microwave radiation offers a new means of studying air showers at E≥10^{17} eV.

Microsatellite instability and Beta2-Microglobulin mutations as prognostic markers in colon cancer: results of the FOGT-4 trial.

BACKGROUND: High-level microsatellite instability (MSI-H) has been reported as a prognostic marker in colon cancer. We here analysed the prognostic significance of MSI and mutations of the Beta2-Microglobulin (B2M) gene, which occur in about 30% of MSI-H colon cancer, in the cohort of the prospective FOGT-4 (Forschungsruppe Onkologie Gastrointestinale Tumoren, FOGT) trial. METHODS: Microsatellite instability status was determined using standard protocols (NCI/ICG-HNPCC panel and CAT25) in 223 colon cancer lesions. Beta2-Microglobulin mutation status was evaluated by exon-wise sequencing in all MSI-H lesions. RESULTS: Patients with MSI-H (n=34) colon cancer presented with a significantly lower risk of relapse after 12 months of follow-up compared with MSS (n=189) colon cancer patients (5 year time to relapse: MSI-H 0.82 vs MSS 0.66, P=0.03). No significant difference in overall survival was detected. Beta2-Microglobulin mutations were identified in 10 (29.4%) out of 34 MSI-H colon cancers and were associated with a complete absence of disease relapse or tumour-related death events (P=0.09). CONCLUSION: The risk of late disease relapse was significantly lower in patients with MSI-H compared with MSS colon cancer. Moreover, B2M mutations may contribute to the favourable outcome of MSI-H colon cancer patients and should therefore be evaluated as a potential prognostic marker in future clinical trials.

Kneelike structure in the spectrum of the heavy component of cosmic rays observed with KASCADE-Grande.

We report the observation of a steepening in the cosmic ray energy spectrum of heavy primary particles at about 8×10(16) eV. This structure is also seen in the all-particle energy spectrum, but is less significant. Whereas the "knee" of the cosmic ray spectrum at 3-5×10(15) eV was assigned to light primary masses by the KASCADE experiment, the new structure found by the KASCADE-Grande experiment is caused by heavy primaries. The result is obtained by independent measurements of the charged particle and muon components of the secondary particles of extensive air showers in the primary energy range of 10(16) to 10(18) eV. The data are analyzed on a single-event basis taking into account also the correlation of the two observables.

[Multimodal therapy for colon cancer: state of the art]. / Kolonkarzinom: Aktueller Stand der multimodalen Therapie.

In UICC stage I a selected group of patients with T1 tumours and a low risk profile regarding simultaneous lymph node metastases can be treated by endoscopic resection alone, if the tumour is thereby completely removed. In UICC stage II an adjuvant chemotherapy (CT) should not be routinely performed. However, in high risk UICC stage II patients (T4 tumour, less than 12 examined lymph nodes, emergency surgery, intraoperative tumour perforation), an adjuvant CT with infusional 5-FU/FA should be recommended. The state of the art in UICC stage III is an adjuvant CT with FOLFOX. In this tumour stage no beneficial effect of CT involving irinotecan or monoclonal antibodies has been documented. Due to CT-induced side effects an infusional 5-FU/FA protocol or oral capecitabine should be given in patients older than 70 years. In stage UICC IV with resectable liver metastases, surgical resection of the primary tumour and the metastases should be implemented. Since no conclusive data are currently available regarding the beneficial effect of neoadjuvant, perioperative or adjuvant CT in this setting, the therapeutic strategy should be individually discussed between surgeons and oncologists (tumour board). In cases of non-resectable liver metastases a neoadjuvant CT should be performed, preferentially with a FOLFOX protocol in combination with targeted therapies, i.e., the monoclonal antibody cetuximab, aimed at tumour regression with radical metastasectomy as the secondary intent (R0). Patients with UICC stage II colon cancer and microsatellite instability (MSI) apparently experience a better prognosis but do not profit from an adjuvant CT with 5-FU/FA alone. If a CT is under consideration for these patients, the MSI status should be determined on tumour tissue. In cases of a positive result a combination CT, i.e., with FOLFOX, should be given. The relevance of the MSI status in other tumour stages is as yet unknown. Before targeted therapies, i.e., cetuximab or panitumumab, are initiated, the KRAS status needs to be determined, since therapies with antibodies against the epithelial growth factor receptor (EGFR) are only effective in tumours bearing the KRAS wild-type.

Adjuvant chemoradiotherapy of advanced resectable rectal cancer: results of a randomised trial comparing modulation of 5-fluorouracil with folinic acid or with interferon-α.

BACKGROUND: Standard adjuvant chemoradiotherapy of rectal cancer still consists of 5-fluorouracil (5-FU) only. Its cytotoxicity is enhanced by folinic acid (FA) and interferon-α (INFα). In this trial, the effects of FA and IFNα on adjuvant 5-FU chemoradiotherapy in locally advanced rectal cancer were investigated. METHODS: Patients with R(0)-resected rectal cancer (UICC stage II and III) were stratified and randomised to a 12-month adjuvant chemoradiotherapy with 5-FU, 5-FU+FA, or 5-FU+IFNα. All patients received levamisol and local irradiation with 50.4 Gy. RESULTS: Median follow-up was 4.9 years (n=796). Toxicities (WHO III+IV) were observed in 32, 28, and 58% of patients receiving 5-FU, 5-FU+FA, and 5-FU+IFNα, respectively. No differences between the groups were observed for local or distant recurrence. Five-year overall survival (OS) rates were 60.3% (95% confidence interval (CI): 54.3-65.8), 60.4% (54.4-65.8), and 59.9% (53.0-66.1) for 5-FU, 5-FU+FA, and 5-FU+IFNα, respectively. A subgroup analysis in stage II (pT3/4pN0) disease (n=271) revealed that the addition of FA tended to reduce the 5-year local recurrence (LR) rate by 55% and increase recurrence-free survival and OS rates by 12 and 13%, respectively, relative to 5-FU alone. CONCLUSIONS: Interferon-α cannot be recommended for adjuvant chemoradiotherapy of rectal cancer. In UICC stage II disease, the addition of FA tended to lower LR and increased survival. The addition of FA to 5-FU may be an effective option for adjuvant chemoradiotherapy of UICC stage II rectal cancer.

GAD65 vaccination: 5 years of follow-up in a randomised dose-escalating study in adult-onset autoimmune diabetes.

AIMS/HYPOTHESIS: The aim of this study was to ascertain whether treatment of GAD65 autoantibody (GADA)-positive diabetic patients with alum-formulated recombinant GAD65 (GAD-alum) is safe and does not compromise beta cell function. METHODS: This Phase 2, placebo-controlled, dose-escalation clinical trial, which was randomized through a central office, was performed in 47 GADA-positive type 2 diabetic patients, who received subcutaneous injections of GAD-alum (4 [n = 9], 20 [n = 8], 100 [n = 9] or 500 [n = 8] microg) or placebo (n = 13) at weeks 1 and 4 of the trial. Participants and caregivers were blinded to group assignments. The primary outcome was safety as assessed by neurological tests, medications and beta cell function evaluated over 5 years, representing the end of the trial. RESULTS: No severe study-related adverse events occurred during the 5 year follow-up. None of the dose groups was associated with an increased risk of starting insulin treatment compared with the placebo group. The use of oral hypoglycaemic agents did not differ between the dose groups. After 5 years, fasting C-peptide levels declined in the placebo group (-0.24; 95% CI -0.41 to -0.07 log(10) nmol/l; p = 0.01) and the 500 microg dose group (-0.37; 95% CI -0.57 to -0.17 log(10) nmol/l; p = 0.003), but not in the 4 microg (-0.10; 95% CI -0.28 to 0.07 log(10) nmol/l; p = 0.20), 20 microg (0.04; 95% CI -0.12 to 0.19 log(10) nmol/l; p = 0.58) and 100 microg (0.00; 95% CI -0.20 to -0.20 log(10) nmol/l; p = 0.98) dose groups. CONCLUSIONS/INTERPRETATION: The primary outcome of safety was achieved, since no severe study-related adverse events occurred. TRIAL REGISTRATION: Because the study was initiated before 1 July 2005, the protocol was not registered in a registry.

Engineering ligand-responsive gene-control elements: lessons learned from natural riboswitches.

In the last two decades, remarkable advances have been made in the development of technologies used to engineer new aptamers and ribozymes. This has encouraged interest among researchers who seek to create new types of gene-control systems that can be made to respond specifically to small-molecule signals. Validation of the fact that RNA molecules can exhibit the characteristics needed to serve as precision genetic switches has come from the discovery of numerous classes of natural ligand-sensing RNAs called riboswitches. Although a great deal of progress has been made toward engineering useful designer riboswitches, considerable advances are needed before the performance characteristics of these RNAs match those of protein systems that have been co-opted to regulate gene expression. In this review, we will evaluate the potential for engineered RNAs to regulate gene expression and lay out possible paths to designer riboswitches based on currently available technologies. Furthermore, we will discuss some technical advances that would empower RNA engineers who seek to make routine the production of designer riboswitches that can function in eukaryotes.

European Academy of Andrology guideline Management of oligo-astheno-teratozoospermia.

BACKGROUND: Oligo-astheno-teratozoospermia is frequently reported in men from infertile couples. Its etiology remains, in the majority of cases, unknown with a variety of factors to contribute to its pathogenesis. The aim of this European Academy of Andrology guideline was to provide an overview of these factors and to discuss available management options. MATERIALS AND METHODS: PubMed was searched for papers in English for articles with search terms: male infertility and oligo-astheno-teratozoospermia. For evidence-based recommendations, the GRADE system was applied. Issues related to urogenital infections/inflammations have not been included in this document as they will be covered by separate guidelines. RESULTS: For men with oligo-astheno-teratozoospermia, the European Academy of Andrology recommends: A general physical examination to assess signs of hypogonadism. A scrotal physical examination to assess (i) the testes and epididymes for volume and consistency, (ii) deferent ducts for total or partial absence, and (iii) occurrence of varicocoele. Performing two semen analyses, according to World Health Organization guidelines to define an oligo-astheno-teratozoospermia. An endocrine evaluation. A scrotal ultrasound as part of routine investigation. Karyotype analysis and assessment of Yq microdeletions in infertile men with a sperm concentration ≤5 × 106 /mL. Cystic fibrosis transmembrane conductance regulator gene evaluation in case of suspicion for incomplete congenital obstruction of the genital tract. Against quitting physical activity to improve the chance of achieving pregnancy. Against androgen replacement therapy to improve the chance of achieving pregnancy. Assisted reproduction techniques to improve the chance of achieving pregnancy, in case other treatment options are not available or not efficient. Androgen replacement therapy in patients with biochemical/clinical signs of hypogonadism, after completion of the fertility treatment. CONCLUSION: These guidelines can be applied in clinical work and indicate future research needs.

Hypogonadism in testicular cancer patients is associated with risk factors of cardiovascular disease and the metabolic syndrome.

More than 95% of testicular cancer are cured but they are at increased long-term risk of cardiovascular disease. The risk of cardiovascular disease and treatment intensity was reported, but it is unknown whether this effect of cancer therapy is direct or indirect, mediated through androgen deficiency. Our aim was, therefore, to evaluate whether testicular cancer patients have increased the prevalence of risk factors of cardiovascular disease and if these risk factors are associated with hypogonadism and/or the cancer treatment given. In 92 testicular cancer patients (mean 9.2 years follow-up) and age-matched controls, blood samples were analysed for lipids, total testosterone, luteinizing hormone (LH), glucose and insulin. An estimate of insulin resistance, HOMAir was calculated. Hypogonadism was defined as total testosterone < 10 nmol/L and/or LH > 10 IU/L and/or androgen replacement. In testicular cancer men with hypogonadism, compared with eugonadal patients, higher insulin (mean difference: 3.10 mIU/L; p = 0.002) and HOMAir (mean difference: 0.792; p = 0.007) were detected. Hypogonadism group presented with increased risk (OR = 4.4; p = 0.01) of metabolic syndrome. Most associations between the treatment given and the metabolic parameters became statistically non-significant after adjustment for hypogonadism. In conclusion, testicular cancer patients with signs of hypogonadism presented with significantly increased risk of metabolic syndrome and investigation of endocrine and metabolic parameters is warranted in these patients.

Prospective correlative chemosensitivity testing in high-dose intraarterial chemotherapy for liver metastases.

Clinical response of liver metastases treated by high-dose intraarterial chemotherapy (HDIAC) delivered via the hepatic artery was predicted by a modification of the human tumor colony-forming assay (HTCFA) originally described by Hamburger and Salmon [Science (Wash. DC), 197:461-463, 1977. In a first set of experiments, the immediate clinical response to HDIAC was determined in 12 patients with colorectal liver metastases. Biopsies were taken immediately before and after HDIAC, and cells were plated in the HTCFA. Three patients received intraoperative 4-epidoxorubicin and another 9 received mitomycin C by 15-min intraarterial infusions. Sensitivity in the HTCFA was defined as 50% inhibition of colony formation in tumors exposed to the chemotherapeutic agent, compared to the untreated controls. Clinical response was accurately predicted by the HTCFA in 11 of 12 cases. Eight patients had a regression of disease following HDIAC treatment with mitomycin C, as evidenced by either greater than 50% reduction in carcinoembryonic antigen serum level (7 patients) or regression of tumor by computed tomography scan (1 patient). Three patients had no evidence of clinical response to epidoxorubicin, and their tumors were resistant to epidoxorubicin in the HTCFA. One tumor was sensitive to mitomycin C in the HTCFA, but serum carcinoembryonic antigen in the patient continued to increase following HDIAC. The HTCFA was also performed on untreated biopsies following incubation in vitro with the drug used for HDIAC. Results correlated with clinical response in all 12 cases. In a second set of experiments, the HTCFA was used to predict the long-term clinical response to HDIAC of 30 patients with liver metastases. One patient had breast cancer metastases, one patient had carcinoid liver metastases, 4 had liver metastases of malignant melanoma, and 24 patients had colorectal liver metastases. All 21 of the patients whose tumors were sensitive in vitro had clinical response, while 6 of 9 patients predicted by the HTCFA to be resistant had no clinical response. Our results demonstrate a high correlation between the HTCFA and clinical response.

Differential expression of CD44 splice variants in human pancreatic adenocarcinoma and in normal pancreas.

Immunohistochemical screening of pancreatic adenocarcinomas from 24 different patients and 9 pancreatic carcinoma cell lines revealed variant CD44 expression in all specimens tested. In contrast to normal pancreatic tissue, carcinomas were strongly positive for epitopes encoded by variant exons v5, whereas v6 was expressed on carcinoma cells as well as normal ductal pancreatic cells. Analysis of RNA expression revealed clear differences between normal pancreatic tissue and tumor specimens. In normal pancreas, v6 and v3 solely and one major chain consisting of v6-v10 were expressed, whereas in pancreatic carcinoma, multiple splice variants were detected. In about 80% of all carcinoma cases and all cell lines tested, the exon v5 appeared in the chain containing at least v4-v10. These data thus far suggest that not the presence alone but the chain composition of the CD44 variant chains could be important for their altered function because one of the major differences between normal and cancer tissue is the linkage of CD44v5 to the CD44v6-containing chain.

UBASH3B/Sts-1-CBL axis regulates myeloid proliferation in human preleukemia induced by AML1-ETO.

The t(8;21) rearrangement, which creates the AML1-ETO fusion protein, represents the most common chromosomal translocation in acute myeloid leukemia (AML). Clinical data suggest that CBL mutations are a frequent event in t(8;21) AML, but the role of CBL in AML1-ETO-induced leukemia has not been investigated. In this study, we demonstrate that CBL mutations collaborate with AML1-ETO to expand human CD34+ cells both in vitro and in a xenograft model. CBL depletion by shRNA also promotes the growth of AML1-ETO cells, demonstrating the inhibitory function of endogenous CBL in t(8;21) AML. Mechanistically, loss of CBL function confers hyper-responsiveness to thrombopoietin and enhances STAT5/AKT/ERK/Src signaling in AML1-ETO cells. Interestingly, we found the protein tyrosine phosphatase UBASH3B/Sts-1, which is known to inhibit CBL function, is upregulated by AML1-ETO through transcriptional and miR-9-mediated regulation. UBASH3B/Sts-1 depletion induces an aberrant pattern of CBL phosphorylation and impairs proliferation in AML1-ETO cells. The growth inhibition caused by UBASH3B/Sts-1 depletion can be rescued by ectopic expression of CBL mutants, suggesting that UBASH3B/Sts-1 supports the growth of AML1-ETO cells partly through modulation of CBL function. Our study reveals a role of CBL in restricting myeloid proliferation of human AML1-ETO-induced leukemia, and identifies UBASH3B/Sts-1 as a potential target for pharmaceutical intervention.

Utility of fast cine magnetic resonance imaging and display for the detection of myocardial ischemia in patients not well suited for second harmonic stress echocardiography.

BACKGROUND: Some patients referred for pharmacological stress testing with transthoracic echocardiography (TTE) are unable to undergo testing owing to poor acoustic windows. Fast cine MRI can be used to assess left ventricular contraction, but its utility for detection of myocardial ischemia in patients poorly suited for echocardiography is unknown. METHODS AND RESULTS: One hundred fifty-three patients (86 men and 67 women aged 30 to 88 years) with poor acoustic windows that prevented adequate second harmonic TTE imaging were consecutively referred for MRI to diagnose inducible myocardial ischemia during intravenous dobutamine and atropine. Diagnostic studies were completed in an average of 53 minutes. No patients experienced myocardial infarction, ventricular fibrillation, exacerbation of congestive heart failure, or death. In patients who underwent computer-assisted quantitative coronary angiography, the sensitivity and specificity for detecting a >50% luminal diameter narrowing were 83% and 83%, respectively. In the 103 patients with a negative MRI examination, the cardiovascular occurrence-free survival rate was 97%. CONCLUSIONS: Fast cine cardiac MRI provides a mechanism to assess left ventricular contraction and diagnose inducible myocardial ischemia in patients not well suited for stress echocardiography.

Assessment of coronary arterial restenosis with phase-contrast magnetic resonance imaging measurements of coronary flow reserve.

BACKGROUND: After successful percutaneous coronary arterial revascularization, 25% to 60% of subjects have restenosis, a recurrent coronary arterial narrowing at the site of the intervention. At present, restenosis is usually detected invasively with contrast coronary angiography. This study was performed to determine if phase-contrast MRI (PC-MRI) could be used to detect restenosis noninvasively in patients with recurrent chest pain after percutaneous revascularization. METHODS AND RESULTS: Seventeen patients (15 men, 2 women, age 36 to 77 years) with recurrent chest pain >3 months after successful percutaneous intervention underwent PC-MRI measurements of coronary artery flow reserve followed by assessments of stenosis severity with computer-assisted quantitative coronary angiography. The intervention was performed in the left anterior descending coronary artery in 15 patients, one of its diagonal branches in 2 patients, and the right coronary artery in 1 patient. A PC-MRI coronary flow reserve value /=70% and >/=50%, respectively. CONCLUSIONS: Assessments of coronary flow reserve with PC-MRI can be used to identify flow-limiting stenoses (luminal diameter narrowings >70%) in patients with recurrent chest pain in the months after a successful percutaneous intervention.

Visualization and functional assessment of proximal and middle left anterior descending coronary stenoses in humans with magnetic resonance imaging.

BACKGROUND: Coronary artery bypass grafting improves survival in patients with >70% luminal diameter narrowing of the 3 major epicardial coronary arteries, particularly if there is involvement of the proximal portion of the left anterior descending (LAD) coronary artery. Measurement of coronary flow reserve can be used to identify functionally important luminal narrowing of the LAD artery. Although magnetic resonance imaging (MRI) has been used to visualize coronary arteries and to measure flow reserve noninvasively, the utility of MRI for detecting significant LAD stenoses is unknown. METHODS AND RESULTS: Thirty subjects (23 men, 7 women, age 36 to 77 years) underwent MRI visualization of the left main and LAD coronary arteries as well as measurement of flow in the proximal, middle, or distal LAD both at rest and after intravenous adenosine (140 microgram/kg per minute). Immediately thereafter, contrast coronary angiography and when feasible, intracoronary Doppler assessments of coronary flow reserve, were performed. There was a statistically significant correlation between MRI assessments of coronary flow reserve and (a) assessments of coronary arterial stenosis severity by quantitative coronary angiography and (b) invasive measurements of coronary flow reserve (P<0.0001 for both). In comparison to computer-assisted quantitative coronary angiography, the sensitivity and specificity of MRI for identifying a stenosis >70% in the distal left main or proximal/middle LAD arteries was 100% and 83%, respectively. CONCLUSIONS: Noninvasive MRI measures of coronary flow reserve correlated well with similar measures obtained with the use of intracoronary Doppler flow wires and predicted significant coronary stenoses (>70%) with a high degree of sensitivity and specificity. MRI-based measurement of coronary flow reserve may prove useful for identification of patients likely to obtain a survival benefit from coronary artery bypass grafting.

Cardiac cycle-dependent changes in aortic area and distensibility are reduced in older patients with isolated diastolic heart failure and correlate with exercise intolerance.

OBJECTIVES: The goal of this study was to determine if cardiac cycle-dependent changes in proximal thoracic aortic area and distensibility are associated with exercise intolerance in elderly patients with diastolic heart failure (DHF). BACKGROUND: Aortic compliance declines substantially with age. We hypothesized that a reduction in cardiac cycle-dependent changes in thoracic aortic area and distensibility (above that which occurs with aging) could be associated with the exercise intolerance that is prominent in elderly diastolic heart failure patients. METHODS: Thirty subjects (20 healthy individuals [10 < 30 years of age and 10 > 60 years of age] and 10 individuals > the age of 60 years with DHF) underwent a magnetic resonance imaging (MRI) study of the heart and proximal thoracic aorta followed within 48 h by maximal exercise ergometry with expired gas analysis. RESULTS: The patients with DHF had higher resting brachial pulse and systolic blood pressure, left ventricular mass, aortic wall thickness and mean aortic flow velocity, and, compared with healthy older subjects, they had a significant reduction in MRI-assessed cardiac cycle-dependent change in aortic area and distensibility (p < 0.0001) that correlated with diminished peak exercise oxygen consumption (r = 0.79). After controlling for age and gender in a multivariate analysis, thoracic aortic distensibility was a significant predictor of peak exercise oxygen consumption (p < 0.04). CONCLUSIONS: Older patients with isolated DHF have reduced cardiac cycle-dependent changes in proximal thoracic aortic area and distensibility (beyond that which occurs with normal aging), and this correlates with and may contribute to their severe exercise intolerance.

Formation of the single-layer beta-sheet of Borrelia burgdorferi OspA in the absence of the C-terminal capping globular domain.

Borrelia outer surface protein A (OspA) contains a unique single-layer beta-sheet that connects N and C-terminal globular domains. This single-layer beta-sheet segment (beta-strands 8-10) is highly stable in solution, although it is exposed to the solvent on both faces of the sheet and thus it does not contain a hydrophobic core. Here, we tested whether interactions with the C-terminal domain are essential for the formation of the single-layer beta-sheet. We characterized the solution structure, dynamics and stability of an OspA fragment corresponding to beta-strands 1-12 (termed OspA[27-163]), which lacks a majority of the C-terminal globular domain. Analyses of NMR chemical shifts and backbone nuclear Overhauser effect (NOE) connectivities showed that OspA[27-163] is folded except the 12th and final beta-strand. (1)H-(15)N heteronuclear NOE measurements and amide H-(2)H exchange revealed that the single-layer beta-sheet in this fragment is more flexible than the corresponding region in full-length OspA. Thermal-denaturation experiments using differential scanning calorimetry and NMR spectroscopy revealed that the N-terminal globular domain in the fragment has a conformational stability similar to that of the same region in the full-length protein, and that the single-layer beta-sheet region also has a modest thermal stability. These results demonstrate that the unique single-layer beta-sheet retains its conformation in the absence of its interactions with the C-terminal domain. This fragment is significantly smaller than the full-length OspA, and thus it is expected to facilitate studies of the folding mechanism of this unusual beta-sheet structure.

Regional chemotherapy directed by individual chemosensitivity testing in vitro: a prospective decision-aiding trial.

A prospective decision-aiding trial was performed to select drugs for regional chemotherapy of various liver tumors (n = 36) by individual drug testing. The drugs were chosen for hepatic artery infusion according to the individual chemosensitivity of tumor biopsies in the human tumor colony-forming assay (HTCA). In vitro HTCA sensitivity correlated with complete response (CR) + partial response (PR) + no change (NC) 93% of the time and with CR + PR 55% of the time. The test sensitivity was 90%, and the specificity was 67% for CR + PR + NC versus progressive disease (PD), whereas the sensitivity and specificity were 89% and 28%, respectively, for CR + PR versus NC + PD. The overall predictive accuracy of the test was 86% for CR + PR + NC versus PD and 58% for CR + PR versus NC + PD. Overall, 83% of this heterogenous patient group with various tumors achieved CR + PR + NC and a 50% clinical response (CR + PR). In vitro-sensitive patients showed a significantly lower intrahepatic progression rate (7% PD) than in vitro-resistant patients (57%; P < 0.05). These results indicate that the HTCA could identify active drugs for individualized hepatic artery infusion, and patients may profit from the use of in vitro-sensitive drugs.

Augmentation of growth hormone secretion during puberty: evidence for a pulse amplitude-modulated phenomenon.

The augmentation of GH secretion that occurs during puberty has been attributed to changes in sex steroid levels that enhance the frequency and amplitude of GH pulses. To investigate the specific GH pulse characteristics responsible for such augmentation we analyzed the serum GH concentration profiles of 10 boys in Tanner stages I-II of sexual development (group A; aged 10 5/12-15 1/12 yr) and compared their GH pulse characteristics with those of 5 boys at Tanner stages IV-V of development (group B; aged 14 8/12-15 1/12 yr). We also reanalyzed previously reported data from 5 prepubertal boys (group C; aged 13 6/12-15 5/12 yr) before and after 10 weeks of treatment with testosterone enanthate (100 mg/4 weeks, im). Using a pulse detection algorithm that constrains the false positive pulse detection rate to less than 5% (Cluster), we found that group B boys had a significantly higher mean serum GH pulse amplitude compared to group A boys (17.1 +/- 2.6 vs. 8.6 +/- 1.7 ng/mL; P = 0.012), but both groups had the same mean GH pulse frequency (group B, 5.4 +/- 0.5 pulses/24 h vs. group A, 5.5 +/- 0.4 pulses/24 h; P greater than 0.05). Similar changes were found in group C boys before and after testosterone therapy; there was no significant change in GH pulse frequency (6.6 +/- 0.9 before vs. 7.6 +/- 0.5 pulses/24 h after treatment; P greater than 0.05), but there was a significant increase in the GH pulse amplitude after therapy (6.8 +/- 1.6 before vs. 15.4 +/- 2.4 ng/mL after treatment; P = 0.04). When the 24-h GH concentration profiles were analyzed using a mathematically distinct method for the estimation of pulse amplitudes, namely the Fourier expansion time series, we confirmed a significant increase in GH pulse amplitude with later stages of puberty and androgen treatment. We conclude that the augmentation in GH secretion that occurs during either spontaneous puberty or exogenous testosterone therapy is an amplitude-modulated phenomenon, relatively independent of changes in pulse frequency. Such an effect may be secondary to the action of sex steroid hormones modulating either the responsivity of somatotrophs to endogenous GH-releasing hormone, the amount of GH-releasing hormone secreted, or the tonic inhibitory tone of somatostatin.