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PURPOSE Early detection of ovarian cancer, the deadliest gynecologic cancer, is crucial for reducing mortality. Current noninvasive risk assessment measures include protein biomarkers in combination with other clinical factors, which vary in their accuracy. Machine learning can be applied to optimizing the combination of these features, leading to more accurate assessment of malignancy. However, the low prevalence of the disease can make rigorous validation of these tests challenging and can result in unbalanced performance. METHODS MIA3G is a deep feedforward neural network for ovarian cancer risk assessment, using seven protein biomarkers along with age and menopausal status as input features. The algorithm was developed on a heterogenous data set of 1,067 serum specimens from women with adnexal masses (prevalence = 31.8%). It was subsequently validated on a cohort almost twice that size (N = 2,000). RESULTS In the analytical validation data set (prevalence = 4.9%), MIA3G demonstrated a sensitivity of 89.8% and a specificity of 84.02%. The positive predictive value was 22.45%, and the negative predictive value was 99.38%. When stratified by cancer type and stage, MIA3G achieved sensitivities of 94.94% for epithelial ovarian cancer, 76.92% for early-stage cancer, and 98.04% for late-stage cancer. CONCLUSION The balanced performance of MIA3G leads to a high sensitivity and high specificity, a combination that may be clinically useful for providers in evaluating the appropriate management strategy for their patients. Limitations of this work include the largely retrospective nature of the data set and the unequal, albeit random, assignment of histologic subtypes between the training and validation data sets. Future directions may include the addition of new biomarkers or other modalities to strengthen the performance of the algorithm.

Objective: To provide a concise review of the new Food and Drug Administration (FDA)-approved antipsychotic, lumateperone, for use in schizophrenia. Data Sources: A literature search of PubMed was performed (January 2000 to May 2020) using the following key terms: lumateperone, Caplyta, and ITI-007. Abstracts from conferences, review articles, clinical trials, and drug monographs were reviewed. Study Selection and Data Extraction: Relevant English-language monographs and studies conducted in humans were considered. Data Synthesis: Lumateperone was FDA approved for the treatment of schizophrenia in December 2019 based on 2 published randomized, double-blind, placebo-controlled trials. Lumateperone’s pharmacology is consistent with that of other second-generation antipsychotics in that it has a higher affinity for the serotonin (5-HT2A) receptors compared with dopamine (D2) receptors but with lower affinities for α-1 and histaminergic receptors. In addition, it serves as a presynaptic dopamine partial agonist, serotonin reuptake inhibitor, and an indirect modulator of glutamatergic systems. Based on the 4-week clinical trials, lumateperone was well tolerated. Most common treatment-emergent adverse events were headache, somnolence, and dizziness. Relevance to Patient Care and Clinical Practice: At this time, lumateperone had a statistically significant reduction in Positive and Negative Syndrome Scale when compared with placebo and was not significantly associated with the extrapyramidal symptoms (EPS) and metabolic adverse effects commonly seen with other antipsychotics. Conclusions: Lumateperone has the potential to benefit individuals with schizophrenia who are intolerant to the EPSs or metabolic adverse effects of other antipsychotics. However, further head-to-head trials with commercially available antipsychotics are still required to assist in establishing its role in treatment.

Cyclin-dependent kinases (CDKs) lie at the heart of eukaryotic cell cycle control, with different cyclin–CDK complexes initiating DNA replication (S-CDKs) and mitosis (M-CDKs)1,2. However, the principles on which cyclin–CDK complexes organize the temporal order of cell cycle events are contentious3. One model proposes that S-CDKs and M-CDKs are functionally specialized, with substantially different substrate specificities to execute different cell cycle events4–6. A second model proposes that S-CDKs and M-CDKs are redundant with each other, with both acting as sources of overall CDK activity7,8. In this model, increasing CDK activity, rather than CDK substrate specificity, orders cell cycle events9,10. Here we reconcile these two views of core cell cycle control. Using phosphoproteomic assays of in vivo CDK activity in fission yeast, we find that S-CDK and M-CDK substrate specificities are remarkably similar, showing that S-CDKs and M-CDKs are not completely specialized for S phase and mitosis alone. Normally, S-CDK cannot drive mitosis but can do so when protein phosphatase 1 is removed from the centrosome. Thus, increasing S-CDK activity in vivo is sufficient to overcome substrate specificity differences between S-CDK and M-CDK, and allows S-CDK to carry out M-CDK function. Therefore, we unite the two opposing views of cell cycle control, showing that the core cell cycle engine is largely based on a quantitative increase in CDK activity through the cell cycle, combined with minor and surmountable qualitative differences in catalytic specialization of S-CDKs and M-CDKs.

Summary A defining feature of cellular growth is that protein and mRNA amounts scale with cell size so that concentrations remain approximately constant, thereby ensuring similar reaction rates and efficient biosynthesis. A key component of this biosynthetic scaling is the scaling of mRNA amounts with cell size, which occurs even among cells with the same DNA template copy number. Here, we identify RNA polymerase II as a major limiting factor increasing transcription with cell size. Other components of the transcriptional machinery are only minimally limiting and the chromatin environment is largely invariant with size. However, RNA polymerase II activity does not increase in direct proportion to cell size, inconsistent with previously proposed DNA-titration models. Instead, our data support a dynamic equilibrium model where the rate of polymerase loading is proportional to the unengaged nucleoplasmic polymerase concentration. This sublinear transcriptional increase is then balanced by a compensatory increase in mRNA stability as cells get larger. Taken together, our results show how limiting RNA polymerase II and feedback on mRNA stability work in concert to ensure the precise scaling of mRNA amounts across the physiological cell size range.

e17553 Background: The baseline risk for ovarian cancer in the general population is 1.2%1. However, women with germline variants in ovarian cancer-associated genes can have a much higher baseline risk for ovarian cancer, up to 54%2. Such a difference in baseline risk presents a triaging challenging for physicians when evaluating adnexal masses. It is well known that identifying pathogenic variants in hereditary cancer syndrome genes often leads to changes in care. Less is known about the utility of negative genetic testing results, specifically in the context of adnexal mass assessment. Methods: An observational, prospective study was designed to examine the clinical impact of multivariate index assay (MIA) and germline genetic testing in patients with a known adnexal mass planned for surgery. Inclusion criteria were: female subjects aged 18-65, presence of an adnexal mass, receiving MIA, and consideration of germline genetic testing. Enrolled physicians selected patients under their care that met inclusion criteria and filled out a short survey describing how the results of each test impacted clinical management. This study was considered IRB-exempt as all information was de-identified. Results: 23 patients were enrolled that received both MIA and germline genetic testing. 21 cases had negative genetic testing and two had positive genetic testing. Both patients with positive genetic testing had low-risk MIA results. Both had risk-reduction surgery with benign findings. Three patients had high-risk MIA results with negative genetic results. All patients had a serial monitoring plan of care. Nine patients with low-risk MIA results and negative genetic results proceeded to surgery. No malignancies were reported. Of the 21 patients with negative genetic results, 17 were not immediately planned for surgery. Conclusions: While not reaching statistical significance, there is a strong association between a negative genetic result and non-immediate surgical intervention. While positive genetic testing results may lead to more aggressive, prophylactic surgical management, negative genetic results may support an expectant management approach. Effectively ruling out the most common hereditary cancer predisposition syndromes may provide physicians with useful insight into a patient's baseline risk. [Table: see text]

Introduction: It is unclear how the interventions designed to restrict community and in-hospital exposure to the SARS-CoV-2 virus affected the care for stroke patients seeking acute treatment. The objective of the following study was to determine the impact COVID-19 has had on the treatment times for patients evaluated as acute stroke alerts at Baptist Hospital of Miami (BHM). A co-primary objective of the study was to assess the risk of contracting SARS-CoV-2 within 2 weeks from hospital discharge. Methods: This retrospective, two phase study was conducted between December 2019 and April 2020. In phase one, we assessed time from symptom onset to hospital arrival, number of strokes with witnessed onset, and in-hospital treatment times pre & post implementation of Covid-19 preventive exposure measures. In phase two of the study, a telephone survey was conducted on the post implementation group to assess the risk of patients developing symptoms or testing positive for SARS-CoV-2 from hospital admission up to two weeks post discharge. Results: Phase I demonstrated there was a 40% decline in stroke volume, but no significant delay to seek medical attention post implementation of the SARS prevention strategies. On average individuals in the pre-group (n=155) waited approximately 260 minutes (SE=24) to seek medical attention vs. 203 minutes (SE=27) minutes for the post-group (n=87). However, there was nearly a six-fold increase in the percentage of cases with unknown symptom onset post implementation of COVID-19 safety precautions. There was significant delay in administering IV alteplase, increasing from 24 mins (n=16) to 33 mins (n=21) post implementation; delays observed for endovascular treatment were not significant (pre, n=13 mean= 73 mins, post n=12 mean= 82 mins). The volume of patients treated with either IV alteplase and/or endovascular treatment remained similar. Phase II of the study is on-going, results will be available for the ISC. Discussion: The COVID-19 crisis in our community was associated with a six-fold increase in the percentage of cases with unknown stroke onset time. Besides a marked decrease in stroke volume, we did not evidence significant delays to either seek or provide acute stroke care outside a modest increase in door to needle time.

Highlights • Opicapone is a 3rd generation catechol-O-methyl transferase (COMT) inhibitor approved in April 2020 by the FDA. • Opicapone is used once daily as adjunctive to levodopa/carbidopa to reduce “off” periods in patients with Parkinson’s disease. • In clinical trials BIPARK I and II, 50 mg of opicapone once daily was shown to be noninferior to entacapone and reduced the mean off time by about 50 min when compared to placebo. • Most common treatment-emergent adverse events for opicapone were dyskinesia, falls, insomnia, and elevated blood creatine phosphokinase levels; it has not been associated with severe hepatic impairment., Objective: To provide a drug review of the newly FDA approved catechol-O-methyl transferase (COMT) inhibitor, opicapone, for the use of end-of-motor motor fluctuation in adults with Parkinson’s disease. Data sources: A literature search of Pubmed was performed till May 2020 using the following key terms: opicapone, Ongentys, and BIA 9-1067. Review articles, clinical trials, and drug monographs were reviewed. Study selection and data extraction: Relevant English-language monographs and studies conducted in humans were considered. Data synthesis: Opicapone was FDA approved for the treatment of end-of-motor motor fluctuation in adults with Parkinson’s disease in April 2020 based on two published randomized clinical trials that were 14 to 15 weeks in duration called BIPARK I and BIPARK II. Based on the clinical trials, 50 mg of opicapone once daily was shown to be noninferior to entacapone and reduced the mean off time by about 50 min when compared to placebo. Most common treatment-emergent adverse events were dyskinesia, falls, insomnia, and elevated blood creatine phosphokinase levels. Relevance to patient care and clinical practice: Opicapone overcomes the limitations associated with other COMT inhibitors since it is dosed once daily, well tolerated, and has not been associated with the risk of hepatic failure. When switching from entacapone to opicapone a reduction in “off” time of −39.3 min was also seen. Conclusions: Opicapone is a once daily 3rd generation COMT inhibitor that has the potential to benefit patients with Parkinson’s disease who are experiencing end-of-motor fluctuations.

Despite the prime importance of telomeres in chromosome stability, significant mysteries surround the architecture of telomeric chromatin. Through micrococcal nuclease mapping, we show that fission yeast chromosome ends are assembled into distinct protected structures (‘telosomes’) encompassing the telomeric DNA repeats and over half a kilobase of subtelomeric DNA. Telosome formation depends on the conserved telomeric proteins Taz1 and Rap1, and surprisingly, RNA. Although yeast telomeres have long been thought to be free of histones, we show that this is not the case; telomere repeats contain histones. While telomeric histone H3 bears the heterochromatic lys9-methyl mark, we show that this mark is dispensable for telosome formation. Therefore, telomeric chromatin is organized at an architectural level, in which telomere-binding proteins and RNAs impose a unique nucleosome arrangement, and a second level, in which histone modifications are superimposed upon the higher order architecture.

Background It is unclear how interventions designed to restrict community and in-hospital exposure to the SARS-CoV-2 (COVID-19) virus influenced stroke care for patients seeking acute treatment. Therefore, we aimed to determine how these COVID-19 interventions impacted acute stroke treatment times and to assess the risk of contracting COVID-19 due to their stay in our medical center. Methods Retrospective, single center, two-phase study evaluating hospital and community trends from 12/2019 – 04/2020 compared to the previous year and pre/post (n = 156/93) intervention implementation. Phase I assessed stroke treatment times, delay to hospital arrival, and witnessed stroke volume. Phase II, a post-implementation telephone survey, assessed risk of developing symptoms or testing positive for COVID-19. Results Stroke volume declined by 29% (p

Cellular growth control is important for all living organisms, but experimental investigation into this problem is difficult because of the complex range of growth regulatory mechanisms. Here, we have used the fission yeast Schizosaccharomyces pombe to identify potential master regulators of growth. At the restrictive temperature, the S. pombe pat1ts mei4Δ strain enters the meiotic developmental program, but arrests in meiotic G2 phase as mei4+ is essential for meiotic progression. These cells do not grow, even in an abundance of nutrients. To identify regulators of growth that can reverse this growth arrest, we introduced an ORFeome plasmid library into the pat1tsmei4Δ strain. Overexpression of eight genes promoted cell growth; two of these were core RNA polymerase subunits, and one was sck2+, an S6 kinase thought to contribute to TORC1 signalling. Sck2 had the greatest effect on cell growth, and we also show that it significantly increases the cellular transcription rate. These findings indicate, for the first time, that global transcriptional control mediated through S6 kinase signalling is central to cellular growth control., Summary: Using a novel system to identify cell growth regulators, we have discovered a number of factors that activate growth in cells in which growth is developmentally switched off.

The dramatic telomerase-dependent overelongation of telomeres in cells lacking Taz1 (ortholog of human TRF1/TRF2) or Rap1 implicates these proteins in restraint of telomerase activity. However, the modes by which these proteins regulate telomerase remain mysterious. Here we show that the mechanisms underlying excessive telomerase activity differ markedly between taz1Δ and rap1Δ strains. Despite allowing elevated telomerase access, rap1Δ telomeres are processed and synthesized in a cell-cycle-constrained manner similar to that of wild-type cells. In contrast, taz1Δ telomeres are processed with little cell-cycle dependency and recruit telomerase over an abnormally wide range of cell-cycle stages. Furthermore, although taz1Δ telomeres experience transient attrition mediated by replication fork stalling, this is balanced not only by temporal expansion of the telomerase activity period, but also by markedly increased recruitment of telomerase and its accessory factor Est1, suggesting that stalled forks generate robust substrates for telomerase.

While telomere repeat-containing non-coding RNA has been identified in a variety of eukaryotes, its biological role is not yet clear. We have identified telomeric transcripts in fission yeast, a model system that combines precise genetic manipulability with telomeres remarkably similar to those of human. Like human and budding yeast, fission yeast harbours a population of telomeric RNA molecules containing G-rich telomeric repeats transcribed from the subtelomere to the telomere. In addition, we detect substantial levels of C-rich telomeric RNA whose appearance is independent of the RNA-dependent RNA polymerase, suggesting that the telomere repeats themselves serve as promoter sites; multiple distinct subtelomeric RNAs are also present. The regulation of these transcripts depends on the telomere-associated proteins Taz1 and Rap1, as deletion of taz1(+) or rap1(+) leads to increased levels of both telomere repeat-containing and subtelomeric transcripts. In contrast, loss of the heterochromatin proteins Swi6 or Clr4 or the telomerase regulator Rif1 results in elevated subtelomeric RNA levels while telomere-repeat containing transcript levels remain repressed. Coupled with the large body of knowledge surrounding the functions of telomeric and heterochromatin factors in fission yeast, these in vivo analyses suggest testable models for the roles of TERRA in telomere function.

Apoptosis is a mechanism employed by multicellular organisms throughout development as a means of eliminating damaged or otherwise unwanted cells. From oogenesis through fertilization and gastrulation, organisms use an array of cell- and tissue-specific mechanisms to regulate the apoptotic program in response to stress or developmental cues. Since cell death regulation is tightly interwoven with cell cycle and checkpoint controls, and embryos of the fly, fish and frog exhibit unique embryonic cell cycle regulation, it is of great interest to understand how early embryos coordinate these cellular functions.

B-lymphocyte-induced maturation protein (Blimp-1) is a transcriptional repressor that is considered to be a master regulator of terminal B-cell development because it is sufficient to trigger differentiation in the BCL(1)-cell model. Transcription of the c-myc gene is repressed by Blimp-1 during B-cell differentiation. In this study, we have explored the mechanism by which Blimp-1 represses transcription by using Gal4-fusion protein assays and assays in which Blimp-1 represses the natural c-myc promoter. The results show that Blimp-1 represses the c-myc promoter by an active mechanism that is independent of the adjacently bound activator YY1. Blimp-1 contains two regions that independently associate with histone deacetylase (HDAC) and endogenous Blimp-1 in nuclear extracts binds in vitro to the c-myc Blimp-1 site in a complex containing HDAC. The functional importance of recruiting HDAC for Blimp-1-dependent repression of c-myc transcription is supported by two experiments. First, the HDAC inhibitor tricostatin A inhibits Blimp-1-dependent repression in cotransfection assays. Second, a chromatin immunoprecipitation assay shows that expression of Blimp-1 causes deacetylation of histone H3 associated with the c-myc promoter, and this deacetylation depends on the Blimp-1 binding site in the c-myc promoter.

Objectives Interstitial cystitis (IC) is a painful, sterile bladder disorder that occurs primarily in women, many of whom also experience allergies with symptoms that worsen perimenstrually. Increased numbers of activated bladder mast cells have recently been implicated in the pathophysiology of IC. These mast cells express high-affinity estrogen receptors and are located close to increased bladder nerves, many of which contain the neuropeptide substance P (SP). We therefore investigated whether the neurotransmitter acetylcholine (ACh) and SP could activate bladder mast cells and whether estradiol could influence this effect. Methods Bladder pieces from male Sprague-Dawley rats were perfused with carbachol (the stable analogue of ACh), SP, or the mast cell secretagogue compound 48/80 (C48/80) with or without preincubation with beta-estradiol. The effect of carbachol was also investigated after pretreatment with the muscarinic antagonist atropine. Mast cell activation was assessed by release of 3 H-serotonin and morphologic evidence of secretion by light and electron microscopy. Results Carbachol triggered rat bladder mast cell serotonin release in a dose-dependent manner, an effect increased by tissue pretreatment with estradiol and blocked by atropine. The effect of carbachol was accompanied by ultrastructural evidence of mast cell activation and was stronger than that obtained by either C48/ 80 or SP. Conclusions Bladder mast cell activation is neurogenically mediated and augmented by estradiol, findings that could possibly explain the painful symptoms of IC and its prevalence in women, as well as the worsening of symptoms perimenstrually.

Xenopus embryos divide rapidly following fertilization. During this rapid period of cleavage, cell divisions are not sensitive to DNA replication or spindle assembly inhibition. Here, we have investigated the consequences of eliciting DNA damage in these embryos. We show that the rapid cell divisions are not affected by DNA damage. However, as the embryos reach the onset of gastrulation, they undergo rapid and synchronous apoptosis. We have investigated the regulation on this delayed apoptotic response to DNA damage. Next, we have reconstituted a DNA damage cell cycle checkpoint in vitro, demonstrating that all the checkpoint signalling components are present in the embryos but are not activated under the experimental conditions used to generate DNA damage in the embryo.

The growing ageing population is leading to an increase in the number of men presenting with lower urinary tract symptoms. This paper argues in favour of more nurse-led clinics offering a holistic approach to assessing this condition that take account of local needs. New guidelines from the British Association of Urology Nurses are also discussed.

The NHS has moved on from the days when the doctor knew best and chose what to tell, or not tell, a patient. Empowerment is the watchword today, and it is crucial for nurses to explore all means of providing patients with information, from face-to-face interactions, supported by written material, to helping them to seek answers via the world wide web.

The prevention of nonhomologous end‐joining (NHEJ) reactions between chromosome ends is crucial for maintaining genome stability. Although this protective function is known to be fulfilled by a core of conserved telomeric proteins that are collectively known as Shelterin, its mechanistic details remain a mystery. In this issue, Sarthy et al (2009) lend fresh insight by developing an ingenious method to dissect the role of hRAP1 in preventing telomeric NHEJ independently of other Shelterin components.

Rmc1, the cellular receptor for the polytropic class of murine retroviruses, determines the tissue tropism of the virus and therefore plays a critical role in the pathogenesis of polytropic virus-induced leukemia. Previously we reported the physical mapping of this gene to a 5-cM region of mouse chromosome 1 and the construction of a yeast artificial chromosome (YAC) contig across this region. In this report we describe the refinement of the Rmc1 candidate region to approximately 600 kb and the generation of an integrated somatic cell hybrid, YAC, and bacterial artificial chromosome contig spanning the region. A number of genes and loci were physically ordered along the chromosome, including a recently identified candidate for Rmc1.

Myotonic dystrophy (DM) is the most common form of muscular dystrophy and is caused by expansion of a CTG trinucleotide repeat on human chromosome 19. Patients with DM develop atrioventricular conduction disturbances, the principal cardiac manifestation of this disease. The etiology of the pathophysiological changes observed in DM has yet to be resolved. Haploinsufficiency of myotonic dystrophy protein kinase (DMPK), DM locus-associated homeodomain protein (DMAHP) and/or titration of RNA-binding proteins by expanded CUG sequences have been hypothesized to underlie the multi-system defects observed in DM. Using an in vivo murine electrophysiology study, we show that cardiac conduction is exquisitely sensitive to DMPK gene dosage. DMPK-/- mice develop cardiac conduction defects which include first-, second-, and third-degree atrioventricular (A-V) block. Our results demonstrate that the A-V node and the His-Purkinje regions of the conduction system are specifically compromised by DMPK loss. Importantly, DMPK+/- mice develop first-degree heart block, a conduction defect strikingly similar to that observed in DM patients. These results demonstrate that DMPK dosage is a critical element modulating cardiac conduction integrity and conclusively link haploinsufficiency of DMPK with cardiac disease in myotonic dystrophy.

We previously reported the induction of dysplasia, a putative precursor of carcinoma, in the dorsolateral prostates (DLPs) of Noble rats by the combined administration of testosterone (T) and estradiol-17beta (E2) for 16 weeks. Additionally, we demonstrated growth of the AIT, a DLP-derived, androgen-independent, transplantable solid tumor, in castrated syngeneic hosts. In this investigation, using Northern blot hybridization, radioimmunoassays and radioligand assays, we showed that transforming growth factor-alpha (TGFalpha) and epidermal growth factor receptor (EGFR) were expressed at close to non-detectable levels in the ventral prostates but at low, but measurable, levels in the DLPs of untreated rats. Enhanced expression of this ligand and its receptor was detected in the DLPs harboring dysplasia and marked overexpression of these molecules was noted in the AIT. In contrast, epidermal growth factor (EGF) expression was found to be constitutively expressed, at high levels, in both normal and dysplastic DLPs, but virtually absent in the AIT. Immunohistochemical data suggested that EGF, TGFalpha and EGFR were aprocine secretory products of the normal DLP, with TGFalpha and EGF localized to the supranuclear complexes and EGFR to the apical membranes of epithelial cells. Alterations in immunostaining patterns for TGFalpha and EGFR were exclusively detected in the dysplastic lesions in the DLPs of T + E2-treated rats. Enhanced intracytoplasmic localization for both peptides were found to accompany the loss of cell polarity in dysplastic foci. Strong intracytoplasmic immunostaining for TGFalpha was observed in some AIT cells whilst staining for EGFR was present in the membranes of tumor cells that formed psuedoacini. Taken together, our findings suggest that autocrine mechanisms may play an important role early in the carcinogenic process and that progression to an androgen-independent neoplastic growth may be modulated by this signaling pathway.