Therapeutic applications of germline testing for cancer predisposition genes in Asia in the real world.

BACKGROUND: Germline genetic testing is traditionally carried out in patients suspected with hereditary cancer syndrome for enhanced cancer surveillance and/or preventive strategies, but is increasingly carried out for therapeutic indications. MATERIALS AND METHODS: We conducted a retrospective review of patients who underwent germline genetic testing at our centre to determine the prevalence of actionable pathogenic germline variants (PGV) and their clinical utility. RESULTS: From 2000 to 2022, 1154 cancer patients underwent germline testing, with the majority (945/1154) tested with multi-gene panels. Four hundred and eleven (35.6%) patients harboured a PGV and 334 (81%) were clinically actionable. BRCA1/2 accounted for 62.3% of actionable mutations, followed by mismatch repair (18%), and other homologous recombination repair (HRR) genes (19.7%). One hundred and fifty-two germline-positive patients have advanced cancers, and 79 received germline-directed therapies (poly ADP ribose polymerase inhibitors = 75; immunotherapy = 4). Median duration of immunotherapy and poly ADP ribose polymerase were 20.5 months (range 5-40 months) and 8 months (range 1-76 months), respectively. Among BRCA/HRR mutation carriers who received platinum-based chemotherapy, pathological complete response rate in the neoadjuvant setting was 53% (n = 17 breast cancers) and objective response rate was >80% in the advanced setting (n = 71). CONCLUSIONS: One-third of cancer patients tested carried a PGV and ∼80% were clinically actionable. Three-quarters of germline-positive advanced cancer patients received germline-directed therapies in the real world, underscoring the practical utility of germline testing to guide cancer therapeutics.

Learning whom not to trust across early and middle adolescence: A longitudinal neuroimaging study to trusting behavior involving an uncooperative other.

Longitudinal changes in trusting behavior across adolescence and their neural correlates were examined. Neural regions of interest (ROIs) included the medial prefrontal cortex (mPFC), dorsal anterior cingulate cortex (dACC), left anterior insula (AI), bilateral ventral striatum (VS), and right dorsal striatum (DS). Participants (wave 1 age: M = 12.90) played the investor in a Trust Game with an uncooperative trustee three times (1-year interval). Analyses included 77 primarily Dutch participants (33 females). Participants decreased their investments with wave. Furthermore, activity was heightened in mPFC, dACC, and DS during investment and repayment, and in right VS (investment) and AI (repayment). Finally, DS activity during repayment increased with wave. These findings highlight early-middle adolescence as an important period for developing sensitivity to uncooperative behavior.

The development of adolescent trust behavior.

Interpersonal trust shows developmental changes during adolescence. The current study used a longitudinal design to examine the development of trust behavior, the presence of gender differences in these developmental trajectories, and the association between individual differences in these developmental trajectories and perspective-taking abilities. The participants played a trust game with a hypothetical trustworthy partner and a trust game with a hypothetical untrustworthy partner in 3 consecutive years (Mage = 12.55 years, Mage = 13.54 years, and Mage = 14.54 years). Concerning the development of trust behavior, the results showed an age-related increase in initial trust behavior and indicated increasingly adaptive trust behavior with age during untrustworthy interactions, whereas no evidence was found for age-related changes in the adaptation of trust during trustworthy interactions. Gender differences were found for the development of initial trust behavior (with boys showing a stronger increase with age than girls), whereas no support was found for the presence of gender differences in the developmental trajectories of adaptive trust behavior during trustworthy and untrustworthy interactions. Furthermore, no evidence was found for perspective-taking abilities to explain individual differences in the development of initial trust behavior or in the development of adaptive trust behavior during trustworthy and untrustworthy interactions. The results provide evidence that, during adolescence initial trust behavior increased with age, more for boys than for girls, and that both boys and girls showed a stronger adaptive response to the untrustworthy partner but not to the trustworthy partner.

Business continuity in blood services: two case studies from events with potentially catastrophic effect on the national provision of blood components.

BACKGROUND AND OBJECTIVES: NHS Blood and Transplant (NHSBT) and the Australian Red Cross Blood Service (ARCBS) are national blood establishments providing blood components to England and North Wales, and Australia, respectively. In 2012, both services experienced potentially catastrophic challenges to key assets. NHSBT suffered a flood that closed the largest blood-manufacturing centre in Europe, whilst ARCBS experienced the failure of a data centre network switch that rendered the national blood management system inaccessible for 42 h. This paper describes both crisis events, including the immediate actions, recovery procedures and lessons learned. MATERIALS AND METHODS: Both incidents triggered emergency response plans. These included hospital reprovisioning and recovery from the incident. Once normal services had been restored, both events were subjected to root cause analysis (RCA) and production of 'lessons learned' reports. RESULTS: In both scenarios, the key enablers of rapid recovery were established emergency plans, clear leadership and the support of a flexible workforce. Product issues to hospitals were unaffected, and there were no abnormal trends in hospital complaints. RCA identified the importance of risk mitigations that require co-operation with external organizations. Reviews of both events identified opportunities to enhance business resilience through prior identification of external risks and improvements to contingency plans, for example by implementing mass messaging to staff and other stakeholders. CONCLUSIONS: Blood establishment emergency plans tend to focus on responding to mass casualty events. However, consolidation of manufacturing to fewer sites combined with a reliance on national IT systems increases the impact of loss of function. Blood services should develop business continuity plans which include prevention of such losses, and the maintenance of services and disaster recovery.

Relationship between disease activity and type 1 interferon- and other cytokine-inducible gene expression in blood in dermatomyositis and polymyositis.

The objective of this study was to evaluate the relationship between blood mRNA, disease activity and treatment effects in a longitudinal study of patients with dermatomyositis (DM) or polymyositis (PM). In all, 24 patients with DM or PM were followed for up to 6 years (mean of 1.9 years) at 2-7 follow-up visits while receiving standard clinical care. Clinical data and blood samples collected at 80 patient visits were used for the analysis of cytokine-induced gene expression for the signaling pathways of type 1 interferon (IFN), tumor necrosis factor-α, IL-1ß, granulocyte-monocyte colony-stimulating factor, IL-10 and IL-13. A type 1 IFN signature score, but not other cytokine signature scores in the blood of patients with DM or PM, correlated highly with disease activity, decreased significantly with immunomodulatory therapies and showed concordant changes with major changes in disease activity. Type 1 IFN signature score in the blood correlates with disease activity in longitudinal follow-up of individual patients with DM or PM. The type 1 IFN-inducible gene transcripts in the blood have potential utility for monitoring disease activity in patients with DM or PM.

Single-cell nonphotochemical hole burning of ovarian surface epithelial carcinoma and normal cells.

Persistent spectral nonphotochemical hole-burning (NPHB) spectroscopy has recently been applied to dye molecules in cells. The sensitivity of NPHB to the nanoenvironment of the probe is well established. It has been shown that NPHB applied to bulk suspensions of cultured human cells can distinguish between normal and cancer cells. Thus, NPHB has potential as a diagnostic cancer tool. For this reason, the methodology is referred to as hole-burning imaging, by analogy with MRI. The optical dephasing time (T(2)) of the dye in hole-burning image replaces the proton T(1) relaxation time in MRI. In addition to the T(2) mode of operation, there are four other modes including measurement of the spectral hole growth kinetics (HGK). Reported here is that the selectivity and sensitivity of NPHB operating in the HGK mode allow for distinction between normal and carcinoma cells at the single-cell level. The ovarian cell lines are ovarian surface epithelial cells with temperature-sensitive large T antigens (analogously normal) and ovarian surface epithelial carcinoma (OV167) cells. The mitochondrial specific dye used was rhodamine 800 (Molecular Probes). This carbocationic dye is highly specific for the outer and inner membranes of mitochondria. In line with the results for bulk suspensions of the two cell lines, the hole-burning efficiency for OV167 cells was found to be significantly higher than that for normal cells. Theoretical analysis of the HGK data leads to the conclusion that the degree of structural heterogeneity for the probe-host configurations in OV167 cells is lower than in the normal cells. Possible reasons for this are given.

Carcinoma and SV40-transfected normal ovarian surface epithelial cell comparison by nonphotochemical hole burning.

Results are presented of nonphotochemical-hole-burning experiments on the mitochondrial specific dye rhodamine 800 incubated with two human ovarian surface epithelial cell lines: OSE(tsT)-14 normal cells and OV167 carcinoma cells. This dye is selective for the plasma and inner membranes of the mitochondria, as shown by confocal microscopy images. Dispersive hole-growth kinetics of zero-phonon holes are analyzed with theoretical fits, indicating that subcellular structural heterogeneity of the carcinoma cell line is lower relative to the analogous normal cell line. Broadening of holes in the presence of an applied electric field (Stark effect) was used to determine the permanent dipole moment change for the S(0)-->S(1) transition in the two cell lines. For the carcinoma cell line, the permanent dipole moment change value is a factor of 1.5 higher than for the normal cell line. It is speculated that this difference may be related to differences in mitochondrial membrane potentials in the two cell lines.

Selective ET(A) antagonists. 5. Discovery and structure-activity relationships of phenoxyphenylacetic acid derivatives.

The fifth paper in this series describes the culmination of our investigations into the development of a potent and selective ET(A) receptor antagonist for the treatment of diseases mediated by ET-1. Receptor site mapping of several ET(A) antagonists prepared previously identified a common cationic binding site which prompted synthesis of phenoxyphenylacetic acid derivative 13a, which showed good in vitro activity (IC(50) 59 nM, rat aortic ET(A)). Optimization of 13a led to the identification of 27b, which exhibited an IC(50) of 4 nM. Although this did not translate into the expected in vivo potency, a compound of comparable in vitro activity, 27a (RPR118031A), showed a far better pharmacokinetic profile and in vivo potency (75 micromol/kg) and was duly proposed and accepted as a development candidate.

Prolactin receptor localization to the area postrema.

The area postrema, which lacks a blood-brain barrier, was examined for the presence of prolactin receptors, which would render it a potential site for vascular prolactin to directly interact with neuronal elements. Using an in vitro autoradiographic technique, frozen sections of New Zealand white rabbit medulla were incubated with radiolabelled ovine prolactin alone (total binding) or radiolabelled ovine prolactin in the presence of excess unlabelled ovine prolactin (non-specific binding). The specificity of the binding was also assessed using excess unlabelled human prolactin or ovine LH. While excess unlabelled ovine and human prolactin caused a statistically significant reduction in radio labeled prolactin binding, unlabelled LH was without effect. Results reveal the presence of specific prolactin binding sites within the area postrema, a previously unknown prolactin target area of the CNS.

Distribution of substance P positive cells and nerve fibers in the rat thymus.

The immune and nervous systems communicate through an array of signalling molecules which includes substance P. This work investigates the anatomical relationship between substance P nerve fibers, receptors, and substance P positive cells in the thymus. Thymuses from rats were frozen or paraformaldehyde fixed. In vitro autoradiography was used to map the distribution of SP receptors. Immunostaining was used to localize SP positive cells and nerve fibers by transmitted light and confocal microscopy. SP receptors exhibited a broader distribution than previously reported, being present throughout the organ with a preferential concentration in the cortico-medullary zone. While SP fibers were frequently associated with the blood vasculature, they were also present throughout the organ independent of blood vessels and were most prominent in the cortico-medullary zone. SP positive cells followed a similar pattern of distribution as the SP fibers and were present as single cells or aggregates of SP positive cells. Confocal microscopy revealed close spatial contact between the SP positive nerve fibers and the SP positive thymic cells. The close spatial relationship between the SP positive thymic cells and SP positive nerve fibers supports the concept of a structural-functional unit between SP nerve fibers and their potential receptor-bearing target cells in the thymus.

Selective endothelin A receptor antagonists. 3. Discovery and structure-activity relationships of a series of 4-phenoxybutanoic acid derivatives.

The third in this series of papers describes our further progress into the discovery of a potent and selective endothelin A (ETA) receptor antagonist for the potential treatment of diseases in which a pathophysiological role for endothelin has been implicated. These include hypertension, ischemic diseases, and atherosclerosis. In earlier publications we have outlined the discovery and structure-activity relations of two moderately potent series of nonpeptide ETA receptor antagonists. In this paper, we describe how a pharmacophore model for ETA receptor binding was developed which enabled these two series of compounds to be merged into a single class of 4-phenoxybutanoic acid derivatives. The subsequent optimization of in vitro activity against the ETA receptor led to the discovery of (R)-4-[2-cyano-5-(3-pyridylmethoxy)phenoxy]-4-(2-methylphenyl)b utanoi c acid (12m). This compound exhibits low-nanomolar binding to the ETA receptor and a greater than 1000-fold selectivity over the ETB receptor. Data are presented to demonstrate that 12m is orally bioavailable in the rat and is a functional antagonist in vitro and in vivo of ET-1-induced vasoconstriction.

Training environment for inferior vena caval filter placement.

This paper describes a Virtual Environment system designed to aid in training interventional radiologists in inferior vena cava filter placement. It is being developed as part of a VE simulator for a number of surgical and interventional radiology procedures at the Laboratory for Advanced Computer Applications in Medicine at the George Washington University. In this procedure a filter is placed in the inferior vena cava to prevent blood clots from the lower portion of the body from reaching the lungs and causing a pulmonary embolus. The simulation is designed to provide both tutorial and testing modes for the filter placement procedure.

New low-density lipoprotein receptor upregulators acting via a novel mechanism.

The synthesis and biological activity of a new series of benzamides and related compounds that upregulate the expression of the low-density lipoprotein (LDL) receptor in human hepatocytes (HepG2 cells) by a novel mechanism are described. The lead compound, N-[5-[(3-cyclohexylpropionyl)amino]-2-methylphenyl]-4-hydroxybe nzamide (1, RPR102359), increased the expression of the LDL receptors in HepG2 cells by 80% when tested at a concentration of 3 microM. Mevinolin (lovastatin) was found to increase the LDL receptor expression by 70% at the same concentration. In contrast to mevinolin, 1 was found to have no effect on cholesterol biosynthesis in liver homogenates or in HepG2 cells at doses where substantial upregulation of the LDL receptor was observed and thus stimulated LDL receptor expression by a novel mechanism.

Distribution of specific substance P binding sites in the heart and adjacent great vessels of the Wistar white rat.

Magnesium(Mg)-deficiency, whether dietary or an effect of a clinical condition such as diabetes, results in a variety of cardiovascular pathologies. Substance P (SP) has been implicated in the induction of cardiac focal inflammatory lesions that occur during Mg-deficiency. Blockade of SP receptors results in a significant reduction in the incidence of lesion formation. In an effort to identify potential endogenous cell populations of the heart, which may play a role in SP-dependent lesion formation, film- and light-microscopic autoradiography were used to map the distribution of specific SP binding sites in frozen sections of the normal rat heart and adjacent great vessels. Binding was assessed with 0.1 nM I-125 Bolton-Hunter labelled SP in the absence (total binding) or presence (non-specific binding) of excess unlabelled SP, prolactin, or L-703,606, a non-peptide antagonist of SP receptors. Film autoradiograms revealed prominent small foci of intense autoradiographic reactions dispersed intermittently around the periphery of the great vessels and coronary arteries, among the interstitial connective tissue of the heart, and along the cusps of the cardiac valves. Excess unlabelled SP caused a significant reduction (97.7% displacement; P < 0.001) in the focal autoradiographic reactions. L-703,606 caused a similar reduction in SP binding (97.3% displacement; P < 0.001), while prolactin had no statistically significant effect on the binding of radiolabelled SP. Light-microscopic autoradiograms revealed that the SP binding sites occurred within clusters of connective tissue cells or in rarely observed parasympathetic ganglia. No evidence was found to suggest the presence of SP receptors on endothelial cells, cardiac muscle fibers, or smooth muscle fibers. The connective tissue cells which bound SP within the heart will likely include types that are susceptible to SP activation and thus may play a role in initiation of the focal inflammation characteristic of Mg-deficiency.

Placental lactogen binding sites in the pregnant rabbit choroid plexus.

Prolactin has direct effects on the CNS. The highest concentration of prolactin receptors resides within the choroid plexus where they probably function to transport prolactin from blood into CSF. Another member of the lactogen family of hormones, placental lactogen (PL), also affects CNS activity and may similarly employ the cerebroventricular system as an intermediary. In order to determine whether the choroid plexus was a PL target tissue, in vitro autoradiography was used to identify specific PL binding sites in the choroid plexus of pregnant New Zealand White rabbits. Frozen brain sections were incubated in a medium containing 125I human PL (hPL) alone (total binding) or with a 500-fold excess of unlabelled hPL (nonspecific binding). The specificity of the binding was assessed with unlabelled human growth hormone (hGH) and ovine luteinising hormone (oLH). An intense autoradiographic reaction occurred over the choroid plexus of tissue sections incubated with 125I hPL alone. Excess unlabelled hPL and hGH, which is lactogenic in the rabbit, caused a significant reduction (P < 0.001) in the binding of radiolabelled hPL to the choroid plexus. In contrast, unlabelled oLH had no effect on radiolabelled hPL binding to this tissue. The results support a role for the choroid plexus in the interactions between PL and the CNS.

Syntheses and biological activities of potent potassium channel openers derived from (+/-)-2-oxo-1-pyridin-3-yl-cyclohexanecarbothioic acid methylamide: new potassium channel openers.

The syntheses and biological activities of (+/-)-2-(cyanomethylene)-1-pyridin-3-ylcyclohexanecarbothioic++ + acid methylamide (6) and trans-(+/-)-2-(cyanomethyl)-1-pyridin-3-ylcyclohexanecarbothioic acid methylamide (14) derived from (+/-)-2-oxo-1-pyridin-3-ylcyclohexanecarbothioic acid methylamide (4) are reported. Compounds were tested for antagonism of potassium-induced contraction of de-endothelialized rat aorta. The effects of modification of 6 and 14 on in vitro K(+)-channel opening activity are presented. These new series of potassium channel openers so derived are best exemplified by (+/-)-2-[2-(phenylsulfanyl)ethylidene]-1-pyridin-3-ylcyclohexan ecarbothioic acid methylamide (13d, RP 66266) and trans-(+/-)-2-[2-[(phenylsulfonyl)amino]ethyl]-1-pyridin-3- ylcyclohexanecarbothioic acid methylamide (25a, RP 66784), which have IC90 values of 3 and 0.3 nM, respectively. The potency of the most active compounds indicates a possible interaction at an extra binding site. The compounds described herein are potential antihypertensive and antianginal agents.

Synthesis and biological activity of trans(+-)-N-methyl-2-(3-pyridyl)-2-tetrahydrothiopyrancarbothioamide 1-oxide (RP 49356) and analogues: a new class of potassium channel opener.

The synthesis and biological activity of trans-(+-)-N-methyl-2-(3-pyridyl)-2-tetrahydrothiopyrancarbothioamid+ ++ e 1-oxide (8a, RP 49356) and analogues is reported. These compounds constitute a new structural class of K(+)-channel opener. The effects of changes in pyridyl group, thioamide, and thiane ring on in vitro K(+)-channel opening reactivity are discussed. A 3-pyridyl or 3-quinolyl group, a small N-alkyl thioamide function, and a thiane oxide ring, in which the sulfoxide is in a trans relationship to the thioamide, are preferred for activity. Selected compounds were tested intravenously in the normotensive anaesthetized rat for hypotensive effects, and the activities reflect their in vitro K(+)-channel opening activity. This led to further evaluation of compound 8a and the selection of the (-)-enantiomer 8b (RP 52891) for development as an antihypertensive and antianginal agent.

Prolactin enhancement of its own uptake at the choroid plexus.

The choroid plexus contains PRL receptors that function in part to transport PRL from the blood into the cerebrospinal fluid (CSF). The blood PRL concentration of female rats was altered by 1) three daily injections of haloperidol (chronic hyperprolactinemia) with or without bromocriptine administration 4 h before death, 2) bromocriptine alone for 4 h (acute hypoprolactinemia), and 3) a single vascular injection of ovine PRL (acute hyperprolactinemia). Changes in the uptake of PRL by the choroid plexus was assessed by quantitative in vivo autoradiography after the injection of radiolabeled PRL. Correlation of changes in PRL uptake at the choroid plexus with changes in PRL transport from blood to CSF was evaluated by subjecting CSF samples to sodium dodecyl sulfate-polyacrylamide gel electrophoresis after vascular injection of radiolabeled PRL. Autoradiography revealed that both chronic and acute hyperprolactinemia resulted in a significant increase in the uptake of radiolabeled PRL by the choroid plexus compared to that in untreated control animals. In contrast, bromocriptine had no effect on PRL uptake at the choroid plexus relative to that in control (untreated) animals. Chronic hyperprolactinemia, but not acute hyperprolactinemia, resulted in a significant increase in the transport of radiolabeled PRL from the blood to the CSF compared to that in untreated controls. The results are consistent with the up-regulation of PRL receptors in the choroid plexus by circulating PRL and the consequent augmentation of transport of PRL from blood to CSF.

Computer-assisted instructions: a role in teaching human gross anatomy.

Eight computer-assisted instructions were developed concerning selected topics in human gross anatomy. The computer lessons were designed to be very flexible in terms of the selection of material to be presented and make extensive use of colour graphics to explain the anatomy. Voluntary testing with immediate and cumulative feedback is incorporated into the lessons so that the users can evaluate their own progress in mastering the subject material. In an effort to assess the value of the computer lessons, the programs were provided to a volunteer group of 48 first-year medical students from a class of 151 students. At the completion of the gross anatomy course, the student users were requested to complete an anonymous questionnaire regarding their impressions of the value of the computer lessons. In addition, test performance on multiple choice examinations was compared between the users of the computer-assisted instructions and their non-user classmates. The responses in the questionnaires revealed a very positive attitude regarding the value and usefulness of the computer-assisted instructions in learning human gross anatomy. The overall rating of the programs on a scale of 1.0 to 10.0 was 1.8 +/- 1.0 with 1.0 representing 'extremely helpful' and 10.0 being 'of no value'. A comparison of test scores showed no significant difference in test performance between the users of the computer-assisted instructions and the non-users. The results of the study suggest that while the computer lessons provide neither an advantage nor a disadvantage in test performance as evaluated by a multiple choice examination, students perceive the computer-assisted instructions as valuable educational tools in mastering the subject of human gross anatomy. The potential role of the computer-assisted instructions in curriculum development is discussed.

The distribution of lactogen receptors in the mammalian hypothalamus: an in vitro autoradiographic analysis of the rabbit and rat.

The hypothalamus contains a high concentration of lactogen receptors as detected with in vitro radioreceptor assay techniques. In an effort to define the location of the lactogen receptors relative to specific hypothalamic nuclei, an in vitro autoradiography technique was applied to frozen sections of rat and rabbit brains. Three lactogenic hormones, i.e. human growth hormone (hGH), ovine prolactin (oPRL), and rat prolactin (rPRL), were radiolabeled with iodine-125. Competition for observed binding sites was assessed with unlabeled hGH, oPRL, and bovine growth hormone (bGH). Analysis of the autoradiographs with a microcomputer-based densitometry system revealed that the rabbit hypothalamus contains specific lactogen binding sites within the supraoptic, paraventricular, suprachiasmatic, ventromedial, arcuate, and dorsomedial nuclei and the medial preoptic area. Unlabeled bGH was effective in competing for binding sites in all areas when hGH but not oPRL was used as the radiolabeled ligand, suggesting the presence of growth hormone receptors in the rabbit hypothalamus with a distribution similar to that of the lactogen binding sites. In contrast to the rabbit, no lactogen binding sites were detected in the rat hypothalamus regardless of the ligand used in the assay. All of the ligands were successful, however, in detecting lactogen receptors within the rat choroid plexus and liver. The results from the rabbits indicate that the influences of prolactin on hypothalamic activity are mediated via lactogen receptors that are widely distributed throughout the various pertinent hypothalamic nuclei. The broad distribution of lactogen receptors in the rabbit hypothalamus attests to the extensive influence of prolactin on hypothalamic regulatory systems. The results from the rat raise questions as to the nature of rat brain prolactin receptors in comparison to prolactin receptors in rat peripheral tissues.