Upregulated pexophagy limits the capacity of selective autophagy.

Selective autophagy is an essential process to maintain cellular homeostasis through the constant recycling of damaged or superfluous components. Over a dozen selective autophagy pathways mediate the degradation of diverse cellular substrates, but whether these pathways can influence one another remains unknown. We address this question using pexophagy, the autophagic degradation of peroxisomes, as a model. We show in cells that upregulated pexophagy impairs the selective autophagy of both mitochondria and protein aggregates by exhausting the autophagy initiation factor, ULK1. We confirm this finding in cell models of the pexophagy-mediated form of Zellweger Spectrum Disorder, a disease characterized by peroxisome dysfunction. Further, we extend the generalizability of limited selective autophagy by determining that increased protein aggregate degradation reciprocally reduces pexophagy using cell models of Parkinson's Disease and Huntington's Disease. Our findings suggest that the degradative capacity of selective autophagy can become limited by an increase in one substrate.

Arf1-PI4KIIIß positive vesicles regulate PI(3)P signaling to facilitate lysosomal tubule fission.

Formation and fission of tubules from autolysosomes, endolysosomes, or phagolysosomes are required for lysosome reformation. However, the mechanisms governing these processes in these different lysosomal organelles are poorly understood. Thus, the role of phosphatidylinositol-4-phosphate (PI(4)P) is unclear as it was shown to promote the formation of tubules from phagolysosomes but was proposed to inhibit tubule formation on autolysosomes because the loss of PI4KIIIß causes extensive lysosomal tubulation. Using super-resolution live-cell imaging, we show that Arf1-PI4KIIIß positive vesicles are recruited to tubule fission sites from autolysosomes, endolysosomes, and phagolysosomes. Moreover, we show that PI(4)P is required to form autolysosomal tubules and that increased lysosomal tubulation caused by loss of PI4KIIIß represents impaired tubule fission. At the site of fission, we propose that Arf1-PI4KIIIß positive vesicles mediate a PI(3)P signal on lysosomes in a process requiring the lipid transfer protein SEC14L2. Our findings indicate that Arf1-PI4KIIIß positive vesicles and their regulation of PI(3)P are critical components of the lysosomal tubule fission machinery.

Fyn and TOM1L1 are recruited to clathrin-coated pits and regulate Akt signaling.

The epidermal growth factor (EGF) receptor (EGFR) controls many aspects of cell physiology. EGF binding to EGFR elicits the membrane recruitment and activation of phosphatidylinositol-3-kinase, leading to Akt phosphorylation and activation. Concomitantly, EGFR is recruited to clathrin-coated pits (CCPs), eventually leading to receptor endocytosis. Previous work uncovered that clathrin, but not receptor endocytosis, is required for EGF-stimulated Akt activation, and that some EGFR signals are enriched in CCPs. Here, we examine how CCPs control EGFR signaling. The signaling adaptor TOM1L1 and the Src-family kinase Fyn are enriched within a subset of CCPs with unique lifetimes and protein composition. Perturbation of TOM1L1 or Fyn impairs EGF-stimulated phosphorylation of Akt2 but not Akt1. EGF stimulation also triggered the TOM1L1- and Fyn-dependent recruitment of the phosphoinositide 5-phosphatase SHIP2 to CCPs. Thus, the recruitment of TOM1L1 and Fyn to a subset of CCPs underlies a role for these structures in the support of EGFR signaling leading to Akt activation.

Glycogen synthase downregulation rescues the amylopectinosis of murine RBCK1 deficiency.

Longer glucan chains tend to precipitate. Glycogen, by far the largest mammalian glucan and the largest molecule in the cytosol with up to 55 000 glucoses, does not, due to a highly regularly branched spherical structure that allows it to be perfused with cytosol. Aberrant construction of glycogen leads it to precipitate, accumulate into polyglucosan bodies that resemble plant starch amylopectin and cause disease. This pathology, amylopectinosis, is caused by mutations in a series of single genes whose functions are under active study toward understanding the mechanisms of proper glycogen construction. Concurrently, we are characterizing the physicochemical particularities of glycogen and polyglucosans associated with each gene. These genes include GBE1, EPM2A and EPM2B, which respectively encode the glycogen branching enzyme, the glycogen phosphatase laforin and the laforin-interacting E3 ubiquitin ligase malin, for which an unequivocal function is not yet known. Mutations in GBE1 cause a motor neuron disease (adult polyglucosan body disease), and mutations in EPM2A or EPM2B a fatal progressive myoclonus epilepsy (Lafora disease). RBCK1 deficiency causes an amylopectinosis with fatal skeletal and cardiac myopathy (polyglucosan body myopathy 1, OMIM# 615895). RBCK1 is a component of the linear ubiquitin chain assembly complex, with unique functions including generating linear ubiquitin chains and ubiquitinating hydroxyl (versus canonical amine) residues, including of glycogen. In a mouse model we now show (i) that the amylopectinosis of RBCK1 deficiency, like in adult polyglucosan body disease and Lafora disease, affects the brain; (ii) that RBCK1 deficiency glycogen, like in adult polyglucosan body disease and Lafora disease, has overlong branches; (iii) that unlike adult polyglucosan body disease but like Lafora disease, RBCK1 deficiency glycogen is hyperphosphorylated; and finally (iv) that unlike laforin-deficient Lafora disease but like malin-deficient Lafora disease, RBCK1 deficiency's glycogen hyperphosphorylation is limited to precipitated polyglucosans. In summary, the fundamental glycogen pathology of RBCK1 deficiency recapitulates that of malin-deficient Lafora disease. Additionally, we uncover sex and genetic background effects in RBCK1 deficiency on organ- and brain-region specific amylopectinoses, and in the brain on consequent neuroinflammation and behavioural deficits. Finally, we exploit the portion of the basic glycogen pathology that is common to adult polyglucosan body disease, both forms of Lafora disease and RBCK1 deficiency, namely overlong branches, to show that a unified approach based on downregulating glycogen synthase, the enzyme that elongates glycogen branches, can rescue all four diseases.

Determinant of Prenatal Diagnostic Testing among Women with Increased Risk of Fetal Aneuploidy and Genetic Disorders.

OBJECTIVE: This study aimed to assess factors that influence patients' decisions in accepting prenatal diagnostic testing following genetic counseling for increased risk of fetal aneuploidy. METHODS: This is a retrospective cohort study of women at increased risk of fetal aneuploidy and genetic disorders who had genetic counseling from January 2012 to December 2016 at a single academic center. Demographics, indications for genetic counseling, and rates of diagnostic testing were collected and compared between those who accepted diagnostic testing and those who chose cell free DNA. The variables were analyzed using Chi-square, Fisher's exact test, and multiple logistic regression. RESULT: Of the 2,373 pregnant women who underwent genetic counseling for increased risk of fetal aneuploidy and genetic disorders during the study period, 321 women had diagnostic testing (13.5%). Women at 35 years and older accepted diagnostic testing more than women younger than 35 years (20.7 vs. 11.5%, p < 0.001). Asian women accepted diagnostic testing at 27.7% more than white, non-Hispanic Black, and Hispanic women at 18.0, 12.1, and 11.7%, respectively, p = 0.002. Number of indications for genetic counseling influenced the likelihood of accepting diagnostic testing. Women with one indication had 11.5% acceptance of diagnostic testing, and with two and three indications, it was 17.0 and 29.2%, respectively. The commonest indication for diagnostic testing was cystic hygroma (risk ratio [RR] = 7.5, 95% confidence interval [CI]: 3.12-8.76 p < 0.001). The relative risk of diagnostic testing for fetuses with shortened long bones, femur and humerus, thickened nuchal fold, echogenic bowel, single umbilical artery, and increased nuchal translucency were 4.0, 3.3, 3.1, 2.7, and 2.7, respectively. Abnormal serum analyte alone was associated with less acceptance of diagnostic testing (RR = 0.8, 95% CI: 0.7-0.96, p = 0.017). CONCLUSION: Age, race, ethnicity, and cumulative number of indications for genetic counseling influenced acceptance of diagnostic testing in at-risk women of fetal aneuploidy and genetic disorders. KEY POINTS: · Genetic counseling.. · Fetal aneuploidy.. · Genetic disorders.. · Prenatal diagnostic testing. Prenatal diagnostic testing in women with increased risk of fetal aneuploidy and genetic disorders..

Maintaining social contacts: The physiological relevance of organelle interactions.

Membrane-bound organelles in eukaryotic cells form an interactive network to coordinate and facilitate cellular functions. The formation of close contacts, termed "membrane contact sites" (MCSs), represents an intriguing strategy for organelle interaction and coordinated interplay. Emerging research is rapidly revealing new details of MCSs. They represent ubiquitous and diverse structures, which are important for many aspects of cell physiology and homeostasis. Here, we provide a comprehensive overview of the physiological relevance of organelle contacts. We focus on mitochondria, peroxisomes, the Golgi complex and the plasma membrane, and discuss the most recent findings on their interactions with other subcellular organelles and their multiple functions, including membrane contacts with the ER, lipid droplets and the endosomal/lysosomal compartment.

Longitudinal outcomes with cancer multigene panel testing in previously tested BRCA1/2 negative patients.

Although multigene panel testing (MGPT) is increasingly utilized in clinical practice, there remain limited data on patient-reported outcomes. BRCA 1/2 negative patients were contacted and offered MGPT. Patients completed pre- and posttest counseling, and surveys assessing cognitive, affective and behavioral outcomes at baseline, postdisclosure and 6 and 12 months. Of 317 eligible BRCA1/2 negative patients who discussed the study with research staff, 249 (79%) enrolled. Decliners were more likely to be older, non-White, and recruited by mail or email. Ninety-five percent of enrolled patients proceeded with MGPT. There were no significant changes in anxiety, depression, cancer specific distress or uncertainty postdisclosure. There were significant but small increases in knowledge, cancer-specific distress and depression at 6-12 months. Uncertainty declined over time. Those with a VUS had significant decreases in uncertainty but also small increases in cancer specific distress at 6 and 12 months. Among those with a positive result, medical management recommendations changed in 26% of cases and 2.6% of all tested. Most BRCA1/2 negative patients have favorable psychosocial outcomes after receipt of MGPT results, although small increases in depression and cancer-specific worry may exist and may vary by result. Medical management changed in few patients.

Research participants' experiences with return of genetic research results and preferences for web-based alternatives.

BACKGROUND: While there is increasing interest in sharing genetic research results with participants, how best to communicate the risks, benefits and limitations of research results remains unclear. METHODS: Participants who received genetic research results answered open and closed-ended questions about their experiences receiving results and interest in and advantages and disadvantages of a web-based alternative to genetic counseling. RESULTS: 107 BRCA1/2 negative women with a personal or family history of breast cancer consented to receive genetic research results and 82% completed survey items about their experience. Most participants reported there was nothing they disliked (74%) or would change (85%) about their predisclosure or disclosure session (78% and 89%). They most frequently reported liking the genetic counselor and learning new information. Only 24% and 26% would not be willing to complete predisclosure counseling or disclosure of results by a web-based alternative, respectively. The most frequently reported advantages included convenience and reduced time. Disadvantages included not being able to ask questions, the risk of misunderstanding and the impersonal nature of the encounter. CONCLUSION: Most participants receiving genetic research results report high satisfaction with telephone genetic counseling, but some may be willing to consider self-directed web alternatives for both predisclosure genetic education and return of results.

A Case of Massive Hepatic Infarction in a Patient with HELLP Syndrome.

Background Hepatic infarction is an exceedingly rare complication of hemolysis, elevated liver enzymes, and low platelets syndrome. Few cases have been described in the medical literature and the true incidence remains unknown. It can lead to fulminant liver failure, liver transplant, or death if not promptly addressed. Case Report A 22-year-old primigravida presented with right upper quadrant and epigastric pain at 28 weeks' gestation. She had severely elevated blood pressures requiring intravenous antihypertensives as well as proteinuria, thrombocytopenia, and mild transaminitis. Within 6 hours of admission, her rapidly rising liver function tests (LFTs) necessitated urgent delivery by primary cesarean section. Her liver enzymes continued to rapidly worsen postoperatively and immediate postpartum computed tomography of the abdomen and pelvis revealed massive hepatic infarction, 11 × 10 × 15 cm, of the right lobe of the liver. Her transaminases peaked at alanine transferase of 2,863 IU/L and aspartate transferase of 2,732 IU/L. She received supportive multidisciplinary intensive care, and LFTs returned to normal by postoperative day 20. Conclusion Hepatic infarction is an extraordinarily rare complication of pre-eclampsia. Early recognition and prompt multidisciplinary management are vital to prevent catastrophic bleeding, hepatic failure, and death.

Risk factors for sexual dysfunction in BRCA mutation carriers after risk-reducing salpingo-oophorectomy.

OBJECTIVE: The aim of the study was to identify risk factors for sexual dysfunction in BRCA mutation carriers who have undergone risk-reducing salpingo-oophorectomy (RRSO). METHODS: A cross-sectional study was performed. BRCA1/2 mutation carriers with and without RRSO were surveyed to determine sexual function (Female Sex Function Index [FSFI]), demographics, medical history, sleep quality, depression, and anxiety scores. Characteristics of patients with the lowest quartile of FSFI scores (<14 ±â€Š8.8) were analyzed to identify risk factors for the most severe phenotype. RESULTS: In the 804 women surveyed, 764 underwent RRSO. Of the 529 (69%) carriers with completed FSFI questionnaires in the RRSO cohort, sexual dysfunction was reported in 77.3%. Poor sleep (P = 0.002), hot flashes (P = 0.002), lack of current systemic hormone therapy (HT) use (P = 0.002), depression (P < 0.001), and anxiety (P = 0.001) were associated with sexual dysfunction. In adjusted analyses, depression (adjusted odds ratio [aOR] 2.4, 95% CI, 1.4-4.1) and hot flashes (aOR 1.9, 95% CI, 1.2-3.0) remained significantly associated with sexual dysfunction. Depression was also a significant risk factor for the most severe degree of sexual dysfunction (OR 2.1, 95% CI, 1.3-3.5) and had the greatest impact on Arousal and Satisfaction domain scores of the FSFI. Current systemic HT use seemed to decrease the risk for sexual dysfunction (aOR 0.6, 95% CI, 0.4-1.0). CONCLUSIONS: Sexual dysfunction is highly prevalent in BRCA mutation carriers after RRSO. Depression seems to be a significant risk factor for sexual dysfunction in this patient population and may be under-recognized and undertreated. Patient and provider education on sexual side effects after surgery and risk factors for sexual dysfunction is necessary to decrease postoperative sexual distress. HT may be associated with improved sexual function after surgery.

Pearls in clinical obstetrics: challenges in anticoagulation in pregnancy.

Women are at increased risk for venous thromboembolism (VTE) during both pregnancy and in the post-partum period. We have conducted a comprehensive literature review of the use of anticoagulation in pregnancy for pregnant women at increased risk for VTE. Multiple factors, including physiologic and pharmacokinetic changes make the treatment and prevention of VTE complicated in pregnancy. Adequate treatment and prevention of VTE in pregnancy is critically important, yet good quality medical studies continue to be lacking. There is a growing amount of data for the use of low molecular weight heparin (LMWH) in pregnant women and this remains the treatment of choice in most indications. For both prophylactic and therapeutic treatments, when LMWHs are chosen, monitoring of antifactor Xa level, although controversial, is advised. Women with prosthetic valve who become pregnant face challenges in regard of type of anticoagulation, dosing and monitoring during pregnancy. Delivery options and peripartum care should be defined with a multidisciplinary approach and taking patient's preference and autonomy in consideration. More high-quality research on this topic is needed to guide the clinical care of this unique population.

Pharmacokinetics of 17 alpha hydroxyprogesterone caproate in singleton pregnancy and its influence of maternal body size measures.

BACKGROUND: Reducing spontaneous preterm deliveries is a worldwide public health priority. Although many interventions have been studied, 1 of the most effective treatments to decrease recurrent preterm birth is the use of weekly 17 alpha hydroxy progesterone caproate. Previous studies on the influence of excessive adipose tissue and obesity on the use of 17 alpha hydroxyprogesterone caproate for the prevention of recurrent spontaneous preterm deliveries have shown conflicting findings. OBJECTIVE: To estimate the pharmacokinetics of weekly17 alpha hydroxyprogesterone caproate in singleton and to evaluate the effect of maternal body size on the pharmacokinetics parameters. STUDY DESIGN: A prospective, open-label, longitudinal design was implemented for this population pharmacokinetic study. Plasma samples and clinical variables were collected in pregnant women between 16 and 36 weeks' gestational age, carrying a singleton pregnancy and receiving 17 alpha hydroxyprogesterone caproate, 250 mg intramuscularly weekly for the prevention of recurrent spontaneous preterm birth. Pharmacokinetics parameters and significant clinical covariates were estimated using mixed effect modeling. Four body size indicators were used in the model to predict pharmacokinetics parameters: lean body weight, total body weight, body mass index, and body surface area. RESULTS: A total of 56 pregnant women, aged 18-44 years with body mass index of 14.5-54.6 kg/m2, provided 114 17 alpha hydroxyprogesterone caproate plasma samples concentration for analysis. A 1-compartment model with first-order absorption satisfactorily described 17 alpha hydroxyprogesterone caproate pharmacokinetics. Compared to other body size indicators, lean body weight best explained intersubject variability. Age, race, and gestational age did not influence 17 alpha hydroxyprogesterone caproate pharmacokinetics. Lean body weight was the best descriptor for the influence of body size on 17 alpha hydroxyprogesterone caproate apparent clearance. Simulations showed that administration of a standard fixed dose of 250 mg intramuscularly produced substantially lower 17 alpha hydroxyprogesterone caproate plasma concentrations in pregnant women with body mass index >30 kg/m2 compared to those with body mass index <30 kg/m2. Conversely, adjustment of the standard dose for differences in total body weight among women resulted in markedly higher 17 alpha hydroxyprogesterone caproate concentrations in women with body mass index >30 kg/m2 compared to women with lower body mass index. Administration of doses adjusted for lean body weight produced nearly identical 117 alpha hydroxyprogesterone caproate plasma concentrations in both the low- and high-body mass index groups. CONCLUSION: Population pharmacokinetics analysis indicates the clearance significantly increases with increasing lean body mass. Higher 17 alpha hydroxyprogesterone caproate doses, adjusted by maternal lean body mass, may be required in patients with a body mass index >30 to achieve equivalent plasma concentrations in pregnant women with a body mass index <30. Adjustment of 17 alpha hydroxyprogesterone caproate doses for lean body weight produces equivalent systemic 17 alpha hydroxyprogesterone caproate exposure in pregnant women regardless of body size.

Cervical cerclage for singleton pregnant patients on vaginal progesterone with progressive cervical shortening.

BACKGROUND: Premature cervical ripening plays a significant role in spontaneous preterm birth. Vaginal progesterone is the recommended treatment in singleton pregnancy with incidental short cervix. There is lack of evidence on whether it is beneficial to reinforce the cervix with cerclage when the cervical length becomes progressively shortened <10 mm while on vaginal progesterone. OBJECTIVE: Our aims are to determine whether cerclage with vaginal progesterone will: (1) reduce the overall spontaneous preterm birth rate, (2) prolong pregnancy latency, and (3) improve neonatal outcomes compared to vaginal progesterone alone. STUDY DESIGN: This was a retrospective cohort study at the University of Illinois at Chicago of all women with singleton pregnancy on vaginal progesterone for incidental short cervix, cervical length <20 mm. Only those with progressive cervical length shortening <10 mm who delivered at the University of Illinois at Chicago from January 2013 through December 2016 were included. The decision to perform cerclage was based on individual physician preference. Demographic data; information on serial cervical length status; medical, obstetric, and social history; cerclage vs no cerclage; and neonatal outcomes were compared. RESULTS: A total of 310 women with incidental short cervix on vaginal progesterone were identified, and of these, 75 had progressive shortening cervical length <10 mm and met inclusion criteria. Among the women with extremely shortened cervical length <10 mm, 36 women (48%) had cervical cerclage plus vaginal progesterone, and 39 women (52%) continued on vaginal progesterone alone. The baseline characteristics, mean cervical length (5.06 vs 5.52 mm), and mean gestational age at diagnosis of extreme short cervix (21.5 vs 21.3 weeks) were similar between women who received cerclage vs those who did not, respectively. The mean gestational age at delivery was significantly greater for those with cerclage (34 weeks and 3 days vs 27 weeks and 2 days; P < .001). The rate of spontaneous preterm birth at <37, 35, 32, 28, and 24 weeks were significantly lower in the cerclage group: 44.1% vs 84.2%, 38.2% vs 81.6%, 23.5% vs 78.9%, 14.7% vs 63.2%, and 11.8% vs 39.5%, respectively. The rate of spontaneous preterm birth <37 weeks remained significant after controlling for confounders (relative risk, 0.11; 95% confidence interval, 0.03-0.41; P < .001). The average pregnancy latency was 14 weeks in the cerclage combined with vaginal progesterone group compared to vaginal progesterone alone group. Neonatal intensive care unit admission and development of respiratory distress syndrome were significantly lower in the cerclage group compared to vaginal progesterone alone group: 13 (36.1%) vs 23 (65.7%) (relative risk, 0.55; 95% confidence interval, 0.34-0.90; P = .018) and 8 (22.2%) vs 17 (43.6%) (relative risk, 0.59; 95% confidence interval, 0.29-0.90; P = .027), respectively. Neonates of women with cerclage were also significantly less likely to develop necrotizing enterocolitis or experience neonatal death. CONCLUSION: Our study showed that cerclage plus vaginal progesterone in women with extremely shortened cervix significantly decreased overall spontaneous preterm birth rates, prolonged pregnancy latency by 2-fold, and decreased the overall neonatal morbidity and mortality.

Returning Individual Genetic Research Results to Research Participants: Uptake and Outcomes Among Patients With Breast Cancer.

PURPOSE: Understanding the outcomes of returning individual genetic research results to participants is critical because some genetic variants are found to be associated with health outcomes and have become available for clinical testing. MATERIALS AND METHODS: BRCA1/2-negative women with early-onset breast cancer, multiple primary cancers, or a family history of breast cancer who participated in a gene discovery cancer registry were offered the opportunity to learn their individual genetic research results of 24 breast cancer susceptibility genes with a genetic counselor after predisclosure genetic counseling. Outcomes included uptake of research results, knowledge, informed choice, psychosocial adjustment, uncertainty, satisfaction, and uptake of clinical confirmation testing. RESULTS: Four hundred two potential participants were contacted. One hundred ninety-four participants (48%) did not respond despite multiple attempts, and 85 participants (21%) actively or passively declined. One hundred seven participants (27%) elected for predisclosure counseling and were more likely to be younger, married, and white. Ninety percent of participants who had predisclosure counseling elected to receive their genetic research results, and 89% made an informed choice. Knowledge increased significantly after predisclosure counseling, and anxiety, intrusive cancer-specific distress, uncertainty, and depression declined significantly after receipt of results. General anxiety and intrusive cancer-specific distress declined significantly for both participants with a positive result and those with a negative result. Sixty-four percent of participants had clinical confirmation testing when recommended, including all participants with a mutation in a high-penetrance gene. CONCLUSION: Uptake of genetic research results may be lower than anticipated by hypothetical reports and small select studies. Participants who elected to receive research results with genetic providers did not experience increases in distress or uncertainty, but not all patients return for confirmation testing.

Reproductive Decision-Making in Women with BRCA1/2 Mutations.

Expanded genetic testing of BRCA mutations has led to identification of more reproductive-aged women who test positive for the mutation which might impact attitudes and decisions about relationships, childbearing and the use of preimplantation genetic diagnosis (PGD) and prenatal diagnosis (PND). A cross-sectional survey was administered to 1081 self-reported BRCA carriers to investigate how knowledge of BRCA status influences these issues. The mean age at BRCA test disclosure was 44 years and 36 % reported a personal history of cancer. Of 163 women who were unpartnered, 21.5 % felt more pressure to get married. Of 284 women whose families were not complete, 41 % reported that carrier status impacted their decision to have biological children. Women with a history of cancer were more likely to report that knowledge of BRCA+ status impacted their decision to have a child (OR 1.8, 95 % CI 1-3.2). Fifty-nine percent thought PGD should be offered to mutation carriers and 55.5 % thought PND should be offered. In conclusion, knowledge of BRCA status impacts attitudes regarding relationships and childbearing, and most carriers believe that PGD and PND should be offered to other carriers. This study suggests that BRCA carriers desire and would benefit from reproductive counseling after test disclosure.

Conflicting Interpretation of Genetic Variants and Cancer Risk by Commercial Laboratories as Assessed by the Prospective Registry of Multiplex Testing.

Purpose Massively parallel sequencing allows simultaneous testing of multiple genes associated with cancer susceptibility. Guidelines are available for variant classification; however, interpretation of these guidelines by laboratories and providers may differ and lead to conflicting reporting and, potentially, to inappropriate medical management. We describe conflicting variant interpretations between Clinical Laboratory Improvement Amendments-approved commercial clinical laboratories, as reported to the Prospective Registry of Multiplex Testing (PROMPT), an online genetic registry. Methods Clinical data and genetic testing results were gathered from 1,191 individuals tested for inherited cancer susceptibility and self-enrolled in PROMPT between September 2014 and October 2015. Overall, 518 participants (603 genetic variants) had a result interpreted by more than one laboratory, including at least one submitted to ClinVar, and these were used as the final cohort for the current analysis. Results Of the 603 variants, 221 (37%) were classified as a variant of uncertain significance (VUS), 191 (32%) as pathogenic, and 34 (6%) as benign. The interpretation differed among reporting laboratories for 155 (26%). Conflicting interpretations were most frequently reported for CHEK2 and ATM, followed by RAD51C, PALB2, BARD1, NBN, and BRIP1. Among all participants, 56 of 518 (11%) had a variant with conflicting interpretations ranging from pathogenic/likely pathogenic to VUS, a discrepancy that may alter medical management. Conclusions Conflicting interpretation of genetic findings from multiplex panel testing used in clinical practice is frequent and may have implications for medical management decisions.

Huntingtin N17 domain is a reactive oxygen species sensor regulating huntingtin phosphorylation and localization.

The N17 domain of the huntingtin protein is post-translationally modified and is the master regulator of huntingtin intracellular localization. In Huntington's disease (HD), mutant huntingtin is hypo-phosphorylated at serines 13 and 16 within N17, and increasing N17 phosphorylation has been shown to be protective in HD mouse models. Thus, N17 phosphorylation is defined as a sub-target of huntingtin for potential therapeutic intervention. We have previously shown that cellular stress can affect huntingtin nuclear entry and phosphorylation. Here, we demonstrate that huntingtin localization can be specifically affected by reactive oxygen species (ROS) stress. We have located the sensor of this stress to the N17 domain, specifically to a highly conserved methionine at position 8. In vitro, we show by circular dichroism spectroscopy structural studies that the alpha-helical structure of N17 changes in response to redox conditions and show that the consequence of this change is enhanced N17 phosphorylation and nuclear targeting of endogenous huntingtin. Using N17 substitution point mutants, we demonstrate that N17 sulphoxidation enhances N17 dissociation from the endoplasmic reticulum (ER) membrane. This enhanced solubility makes N17 a better substrate for phosphorylation and subsequent nuclear retention. This ability of huntingtin to sense ROS levels at the ER, with phosphorylation and nuclear localization as a response, suggests that ROS stress due to aging could be a critical molecular trigger of huntingtin functions and dysfunctions in HD and may explain the age-onset nature of the disorder.

Exercise-Induced Dose-Response Alterations in Adiponectin and Leptin Levels Are Dependent on Body Fat Changes in Women at Risk for Breast Cancer.

BACKGROUND: Dysregulation of adipokines, such as adiponectin and leptin, is associated with a variety of chronic diseases, including cancer. Physical activity protects against breast cancer and one of the mechanisms which may underlie this association is exercise-induced changes in adipokine levels. The WISER Sister Trial was a three-armed randomized controlled trial in premenopausal women (n = 137) with an elevated risk for breast cancer. METHODS: A 5-menstrual-cycle-long dosed aerobic exercise intervention compared low-dose exercise (150 min/wk; n = 44) or high-dose exercise (300 min/wk; n = 48) with a control group asked to maintain usual activity levels (n = 45). Exercise intensity progressed to and was maintained at 70% to 80% of age predicted heart rate max. Body composition and adipokine levels were measured at baseline and follow-up. RESULTS: We observed significant linear trends for increased fitness capacity (Δ%: -2.0% control, 10.1% low dose, 13.1% high dose), decreased fat tissue-to-total tissue mass (Δ%: 0.7% control, -2.9% low dose, -3.7% high dose), increased body fat adjusted adiponectin (Δ%: -0.6% control, 0.6% low dose, 0.9% high dose), and decreased body fat adjusted leptin (Δ%: 0.7% control, -8.2% low dose, -10.2% high dose). CONCLUSIONS: In this randomized clinical trial of premenopausal women at risk for breast cancer, we demonstrate a dose-response effect of exercise on adiponectin and leptin and that dose response is dependent on changes in body fat. IMPACT: Improved adipokine levels, achieved by aerobic exercise training-induced decreases in body fat, may decrease breast cancer risk for high-risk premenopausal women. Cancer Epidemiol Biomarkers Prev; 25(8); 1195-200. ©2016 AACR.

Patient feedback and early outcome data with a novel tiered-binned model for multiplex breast cancer susceptibility testing.

PURPOSE: The risks, benefits, and utilities of multiplex panels for breast cancer susceptibility are unknown, and new counseling and informed consent models are needed. We sought to obtain patient feedback and early outcome data with a novel tiered-binned model for multiplex testing. METHODS: BRCA1/2-negative and untested patients completed pre- and posttest counseling and surveys evaluating testing experiences and cognitive and affective responses to multiplex testing. RESULTS: Of 73 patients, 49 (67%) completed pretest counseling. BRCA1/2-negative patients were more likely to proceed with multiplex testing (86%) than those untested for BRCA1/2 (43%; P < 0.01). Many patients declining testing reported concern for uncertainty and distress. Most patients would not change anything about their pre- (76%) or posttest (89%) counseling sessions. Thirty-three patients (72%) were classified as making an informed choice, including 81% of those who proceeded with multiplex testing. Knowledge increased significantly. Anxiety, depression, uncertainty, and cancer worry did not significantly increase with multiplex testing. CONCLUSION: Some patients, particularly those without prior BRCA1/2 testing, decline multiplex testing. Most patients who proceeded with testing did not experience negative psychological responses, but larger studies are needed. The tiered-binned approach is an innovative genetic counseling and informed consent model for further study in the era of multiplex testing.Genet Med 18 1, 25-33.