Edition 53 - 2015, June / Subject Review

Subject Review: Report of the ENDO2015 97th Annual Meeting of the ENDOCRINE SOCIETY – MARCH 5 – 8, 2015 – Ed. 53

Marco A. Rivarola y Alicia Belgorosky, Servicio de Endocrinologia, Hospital de Pediatria Garrahan, Buenos Aires, Argentina

As announced by the ENDO 2015 Meeting Chairs, “Throughout the program the theme of change will be seen featured in sessions, speakers, and work. This evolution still retains the core of ENDO: cutting edge science and state of the art clinical practice presented by global experts.” In his Welcome remarks, Society President Richard Santen, M.D. told us that for the first time Endocrine Society Meeting membership exceeded 18.000 with 40% of those members coming from outside the United States. This year, he added, International activities were numerous and diverse (Argentina, Brazil, Korea, Turkey, Mexico, Egypt, India, China and Peru). It is recognized that Society members contribute substantial expertise and talent to the Meeting, which serves to elevate the experience of attendees.

PLENARY CONFERENCES

L1-1 Presidential Plenary: Genomics, Pharmacogenomics, and Functional Genomics in Menopausal Women: Implications for Precision Medicine

James N Ingle, MD
Mayo Clinic, Rochester, MN

Talk Description: In postmenopausal women, there is substantial inter-individual variation in plasma estrogens. We have identified a variant SNP, associated with concentrations of estradiol, that creates a functional estrogen response element (ERE). GWAS in women receiving aromatase inhibitors for early breast cancer have identified variant SNPs, associated with musculoskeletal adverse events, that also create an ERE. GWAS in women receiving tamoxifen or raloxifene for breast cancer prevention revealed variant SNPs associated with risk and activity of BRCA1.

The summary of a recent paper by Prof. Ingle follows:

J Hum Genet. 2013 Jun;58(6):306-12.

Ingle JN. Pharmacogenomics of endocrine therapy in breast cancer. 1Division of Medical Oncology, Mayo Clinic, Rochester, MN 55905, USA. Ingle.

Abstract

The most important modality of treatment in the two-thirds of patients with an estrogen receptor (ER)-positive early breast cancer is endocrine therapy. In postmenopausal women, options include the selective ER modulators (SERMs), tamoxifen and raloxifene, and the ‘third-generation’ aromatase inhibitors (AIs), anastrozole, exemestane and letrozole. Under the auspices of the National Institutes of Health Global Alliance for Pharmacogenomics, Japan, the Mayo Clinic Pharmacogenomics Research Network Center and the RIKEN Center for Genomic Medicine have worked collaboratively to perform genome-wide association studies (GWAS) in women treated with both SERMs and AIs. On the basis of the results of the GWAS, scientists at the Mayo Clinic have proceeded with functional genomic laboratory studies. As will be seen in this review, this has led to new knowledge relating to endocrine biology that has provided a clear focus for further research to move toward truly personalized medicine for women with breast cancer.

 

L1-2 Presidential Plenary: Personalized Menopause Management: Clinical and Biomarker Data That Inform Decision Making

JoAnn E Manson, MD, DrPH, FACP, FACE
Brigham and Women’s Hospital, Harvard Medical School, Boston, MA

Talk Description: A critical mass of research now informs the hormonal treatment (HT) decision, but controversy and confusion about HT continue to serve as barriers to appropriate use. The purpose of the conference was to help clinicians and researchers make sense of the evidence, to bring a more rational approach to clinical decision making in this important area. Some, but not all, women are good candidates for HT, and the use of risk stratification and incorporation of the personal preferences of the patient, will guide the approach taken. The array of treatment options, including lower hormone doses and transdermal routes of delivery, has expanded markedly in recent years. A recently released algorithm and mobile app will be discussed, which helps clinicians and patients work together to decide between hormonal and non-hormonal treatment options for menopausal symptoms and develop a personalized treatment plan based on shared decision making.

The summary of a recent paper by Dr. J. E. Manson et al. follows:

Eur J Nutr. 2015 Mar 29.

Wang L1, Manson JE, Rautiainen S, Gaziano JM, Buring JE, Tsai MY, Sesso HD. A prospective study of erythrocyte polyunsaturated fatty acid, weight gain, and risk of becoming overweight or obese in middle-aged and older women. Division of Preventive Medicine, Department of Medicine, Brigham and Women’s Hospital, 900 Commonwealth Avenue East, Boston, MA, 02215, USA,

Abstract

PURPOSE: ω3 and ω6 fatty acids (FA) may have divergent effects on the development of obesity. We examined the association of baseline erythrocyte ω3 and ω6 FA composition with body weight change and the risk of becoming overweight or obese in the Women’s Health Study (WHS) participants. METHODS: We identified 534 women who had baseline erythrocyte FA measured and a baseline body mass index (BMI) of 18.5-<25 kg/m2. Body weight was updated at a total of six time points during follow-up. RESULTS: Weight gain during a mean of 10.4-year follow-up increased with increasing quartiles of baseline erythrocyte cis ω6 FA, ω6/ω3 ratio, and trans FA while decreased with increasing cis ω3 FA. After multivariable adjustment including total energy intake and physical activity, the weight gain (kg) in the highest versus the lowest quartile was 3.08 versus 2.32 for erythrocyte cis ω6 FA (p trend 0.04), 2.07 versus 2.92 for cis ω3 FA (p trend 0.08), 2.93 versus 2.05 for ω6/ω3 ratio (p trend 0.046), and 3.03 versus 2.27 for trans FA (p trend 0.06). Among individual FA, the associations were significant for 18:2ω6, 18:3ω6, and trans 18:1 and marginally significant for 20:3ω6 and trans 18:2. The risk of becoming overweight or obese (defined as BMI ≥25 kg/m2 at any follow-up time point) increased across increasing ω6/ω3 ratio (multivariable model p trend 0.04). CONCLUSIONS: In this prospective study, we found suggestive evidence that erythrocyte cis ω6 FA may be positively associated, and cis ω3 FA inversely associated with weight gain in initially normal-weight women.

L2-1 Neurosteroid Estrogens: Implications for Epilepsy

Session: L2-Thursday Afternoon Plenary
Basic

Catherine S Woolley, PhD
Northwestern University, Evanston, IL

Talk Description: There is growing awareness that estrogens may be produced in the brain as neurosteroids that act locally within specific brain regions to modulate neurophysiology on a time scale of minutes. This lecture will review key findings on acute estrogen actions in the hippocampus, a limbic brain structure commonly involved in temporal lobe epilepsy, and will present pre-clinical evidence of a link between neurosteroid estrogens and the severity of limbic seizures in both sexes.

The summary of a recent paper by Dr. Woolley et al. follows:

Brain Struct Funct. 2014 Nov;219(6):1947-54.

Estradiol regulates large dense core vesicles in the hippocampus of adult female rats. May RM1, Tabatadze N, Czech MM, Woolley CS. Department of Neurobiology, Northwestern University, 2205 Tech Drive, Evanston, IL, 60208, USA.

Abstract

Previous work has shown that the steroid hormone estradiol facilitates the release of anticonvulsant neuropeptides from inhibitory neurons in the hippocampus to suppress seizures. Because neuropeptides are packaged in large dense core vesicles, estradiol may facilitate neuropeptide release through regulation of dense core vesicles. In the current study, we used serial section electron microscopy in the hippocampal CA1 region of adult female rats to test three hypotheses about estradiol regulation of dense core vesicles: (1) Estradiol increases the number of dense core vesicles in axonal boutons, (2) Estradiol increases the size of dense core vesicles in axonal boutons, (3) Estradiol shifts the location of dense core vesicles toward the periphery of axonal boutons, potentially lowering the threshold for neuropeptide release during seizures. We found that estradiol increases the number and size of dense core vesicles in inhibitory axonal boutons, consistent with increased neuropeptide content, but does not shift the location of dense core vesicles closer to the bouton periphery. These effects were specific to large dense core vesicles (>80 nm) in inhibitory boutons. Estradiol had no effects on small dense core vesicles or dense core vesicles in excitatory boutons. Our results indicate that estradiol suppresses seizures at least in part by increasing the potentially releasable pool of neuropeptides in the hippocampus, and that estradiol facilitation of neuropeptide release involves a mechanism other than mobilization of dense core vesicles toward sites of release.

L2-2 Genetics and Pathophysiology of Human Obesity and Metabolism

Session: L2-Thursday Afternoon Plenary
Translational

Stephen O’Rahilly, MD, FRCP
University of Cambridge Metabolic Research Laboratories, Cambridge, United Kingdom

Talk Description:

The genetic component of quantitative metabolic traits is complex with a mixture of common alleles of small effect and rarer alleles of larger effect. We have principally focused on finding the latter through the study of extreme human phenotypes of obesity and insulin resistance, including lipodystrophy. By applying both candidate and hypothesis-free genetic approaches we have identified multiple different genetic variants that cause highly penetrant forms of these diseases.

The summary of a recent book chapter by O´Rahilly S and Farooqi IS follows:

O’Rahilly S, Farooqi IS. The Genetics of Obesity in Humans. In: De Groot LJ, Beck-Peccoz P, Chrousos G, Dungan K, Grossman A, Hershman JM, Koch C, McLachlan R, New M, Rebar R, Singer F, Vinik A, Weickert MO, editors. Endotext [Internet]. South Dartmouth (MA): MDText.com, Inc.; 2000-2013 Mar 23.

Abstract.

Within a population, the variance in body mass index (BMI: weight in kg/height in meters squared) is largely genetically determined. Estimates of heritability (the proportion of the phenotypic variance of a trait that is attributable to genetic variation) range between 40% and 70% based on twin and family studies in a number of populations. In a Swedish study of twins reared apart, within-pair correlations for BMI were 0.70 for male and 0.66 for female monozygotic twins. These observations are supported by more recent studies in twins born after the recent increase in the prevalence of obesity, which have estimated the heritability of BMI at 77%. Whilst the contribution of the shared environment cannot be distinguished in most studies, comparable estimates of heritability have been derived from studies of adopted children, whose weight correlates better with that of their biological parents than with that of their adoptive parents. The genetic contribution to obesity is further supported by population studies of admixture. Genetic factors may modulate the response to changes in energy intake and expenditure as evidenced by overfeeding studies in monozygotic twins which have shown that the weight gain in response to positive energy balance differed across sets of identical twins but was similar between members of a twin pair.

Comments

COMMON VARIANTS IDENTIFIED BY GENOME WIDE ASSOCIATION STUDIES (GWAS)Altogether, GWAS for obesity-related traits (including BMI, waist-hip ratio and obesity) have led to the discovery of at least 52 loci to date. However, the common variants uncovered in GWASs are characterized by modest effect sizes (per-allele odds ratios between 1.2 and 1.5), and the proportion of variability explained by GWAS-identified loci to date remains relatively modest (<5%). Understanding the biology underlying these association signals is still at an early stage pattern of inheritance, however, increasingly more complex models of inheritance are likely to emerge. Severe obesity can result from a multiplicity of defects involving the leptin-melanocortin pathway. Briefly, leptin is an adipocyte-derived hormone whose circulating levels correlate closely with fat mass. Many of the physiological effects of leptin are mediated through the long isoform of the leptin receptor which is widely expressed in the hypothalamus and other brain regions involved in energy homeostasis. Leptin stimulates the expression of pro-opiomelanocortin (POMC) in primary neurons located in the arcuate nucleus of the hypothalamus. POMC is extensively post-translationally modified to generate the melanocortin peptides, which activate the melanocortin receptors to modulate diverse functions in the central nervous system, the adrenal gland, and skin. The melanocortins are agonists at melanocortin receptors and suppress food intake. In addition, leptin inhibits orexigenic pathways, mediated by neurons expressing the melanocortin antagonist Agouti-related protein and neuropeptide Y (NPY); NPY can suppress the expression of POMC. These two sets of primary leptin-responsive neurons project to second-order neurons expressing the melanocortin 4 receptor (MC4R). Targeted genetic disruption of MC4R in mice leads to increased food intake and increased lean mass and linear growth. These hypothalamic pathways interact with other brain centers to coordinate appetite and modulate efferent signals to the periphery, regulating intermediary metabolism and energy expenditure.

  1. OBESITY SYNDROMES

LEPTIN AND LEPTIN RECEPTOR DEFICIENCY Congenital leptin (LEP) and leptin receptor (LEPR) deficiency are rare, autosomal recessive disorders associated with severe obesity from a young age. Homozygous frameshift, nonsense, and missense mutations involving LEP and LEPR have been identified in 1% and 2-3% of severely obese patients from consanguineous families, respectively. Leptin receptor mutations have been found in some non-consanguineous families, where both parents were unrelated but carried rare alleles in heterozygous form.

DISORDERS AFFECTING PRO-OPIOMELANOCORTIN (POMC)-DERIVED PEPTIDES AND POMC PROCESSING. As POMC is a precursor of adrenocorticotrophin (ACTH) in the pituitary, children who are homozygous or compound heterozygous for loss of function mutations in POMC present in neonatal life with hypoglycaemia, cholestatic jaundice or other features of adrenal crisis due to ACTH deficiency. They require long-term corticosteroid replacement therapy. Such children have pale skin, and white Caucasians have red hair, due to the lack of melanocortin function at melanocortin 1 receptors in the skin. Although red hair may be an important diagnostic clue in patients of Caucasian origin, children from different ethnic backgrounds may have a less obvious phenotype such as dark hair with red roots. POMC deficiency results in hyperphagia and early-onset obesity due to loss of melanocortin signaling at the MC4R. The clinical features are comparable to those reported in patients with mutations in the receptor for POMC derived ligands, MC4R.

PCSK1 DEFICIENCY. Many biologically inactive prohormones and neuropeptides are cleaved by serine endoproteases to release biologically active peptides. The prohormone convertase (PCSK1) is expressed in neuroendocrine tissues and acts upon a range of substrates including proinsulin, proglucagon and POMC. Compound heterozygous or homozygous mutations in PCSK1 cause neonatal small bowel enteropathy, glucocorticoid deficiency, hypogonadotropic hypogonadism and postprandial hypoglycaemia due to impaired processing of proinsulin to insulin as well as severe, early onset obesity. Elevated plasma levels of proinsulin and 32–33 split proinsulin in the context of low levels of mature insulin provide the basis for a diagnostic test for this disorder.

MC4R DEFICIENCY. Heterozygous MC4R mutations have been reported in obese people from various ethnic groups. The prevalence of pathogenic MC4R mutations has varied from 0.5 -2.5% of people with a BMI>30 kg/m2 in UK and European populations to 5% in patients with severe childhood obesity. As MC4R deficiency is the most common genetic form of obesity, assessment of the sequence of the MC4R is increasingly seen as a necessary part of the clinical evaluation of the severely obese child. The clinical features of MC4R deficiency include hyperphagia, which often starts in the first few years of life. Alongside the increase in fat mass, MC4R-deficient subjects also have an increase in lean mass and a marked increase in bone mineral density, thus they often appear “big-boned”. They exhibit accelerated linear growth in early childhood, which may be a consequence of disproportionate early hyperinsulinemia and effects on pulsatile growth hormone (GH) secretion, which is retained in MC4R-deficient adults in contrast to common forms of obesity. Despite this early hyperinsulinemia, obese adult subjects who are heterozygous for mutations in the MC4R gene are not at increased risk of developing glucose intolerance and type 2 diabetes compared to controls of similar age and adiposity.

Further studies established that MC4R plays a role in fat oxidation and nutrient partitioning. These effects are also seen in rodents and appear to be mainly mediated by the sympathetic nervous system. Reduced sympathetic nervous system activity in MC4R-deficient patients is likely to explain the lower prevalence of hypertension and lower systolic and diastolic blood pressures. Thus, central melanocortin signalling appears to play an important role in the regulation of blood pressure and its coupling to changes in weight. At present, there is no specific therapy for MC4R deficiency, but patients with heterozygous MC4R mutations do respond to Roux-en-Y-bypass surgery, which can be considered in adults.

ALBRIGHT HEREDITARY OSTEODYSTROPHY. Albright hereditary osteodystrophy (AHO) is an autosomal dominant disorder due to germline mutations in GNAS1 that decrease expression or function of G alpha s protein. Heterozygous loss-of-function mutations lead to AHO, a disease characterized by short stature, obesity, skeletal defects, and impaired olfaction. Maternal transmission of GNAS1 mutations leads to AHO plus resistance to several hormones (e.g., parathyroid hormone) that activate Gs in their target tissues (pseudohypoparathyroidism type IA), while paternal transmission leads only to the AHO phenotype (pseudopseudohypoparathyroidism). Studies in both mice and humans demonstrate that GNAS1 is imprinted in a tissue-specific manner, being expressed primarily from the maternal allele in some tissues and biallelically expressed in most other tissues, thus multi-hormone resistance occurs only when Gs (alpha) mutations are inherited maternally.

SH2B1 DEFICIENCY. Copy number variants (CNVs), which include gains (duplications or insertions), losses (deletions), and rearrangements, have been estimated to account for nearly 18% of the heritable variance in gene expression. We and others have identified several rare CNVs recurrent among severely obese patients and enriched relative to controls. Deletion of a 220-kb segment of 16p11.2 is associated with highly penetrant familial severe early-onset obesity and severe insulin resistance. This deletion includes a small number of genes, one of which is SH2B1 , known to be involved in leptin and insulin signaling. These patients gain weight in the first years of life, with hyperphagia and fasting plasma insulin levels that are disproportionately elevated compared to age- and obesity-matched controls. Several mutations in the SH2B1 gene have also been reported in association with early onset obesity, severe insulin resistance and behavioural abnormalities in some patients.

  1. OBESITY AND DEVELOPMENTAL ABNORMALITIES.

Prader-Willi Syndrome. Prader-Willi syndrome is an autosomal dominant disorder with an estimated prevalence of about 1 in 25,000 caused by deletion or disruption of a paternally imprinted region on chromosome 15. The clinical features of Prader-Willi syndrome (PWS) include diminished foetal activity, hypotonia in infancy, obesity, mental retardation, short stature, hypogonadotropic hypogonadism and small hands and feet. Diagnostic criteria arrived at by a consensus group refer to five major criteria, such as feeding problems in infancy and failure to thrive and seven minor criteria, such as hypopigmentation, which can be added together to give a score which can be used to make a clinical diagnosis of PWS .

Bardet Biedl Syndrome. Bardet Biedl Syndrome. Bardet-Biedl syndrome (BBS) is a rare (prevalence <1>de novo missense mutation impairing the function of TrkB. We also identified a patient with a de novo chromosomal inversion, which encompasses the BDNF locus and disrupts BDNF expression. Yanovski and colleagues showed that in patients with WAGR syndrome, a subset of chromosome 11p.12 deletions encompassing the BDNF locus were associated with early-onset obesity.

SIM1 DEFICIENCY. Single minded 1 (SIM1) is a transcription factor involved in the development of the paraventricular and supraoptic nuclei of the hypothalamus. A de novo balanced translocation between chromosomes 1p22.1 and 6q16.2, which disrupts SIM1 and multiple heterozygous missense mutations in SIM1 cause severe obesity. The Drosophila single-minded (sim) gene is a regulator of fruit fly neurogenesis and in the mouse Sim1 is expressed in the developing kidney and central nervous system and is essential for formation of the supraoptic and paraventricular (PVN) nuclei which express the melanocortin-4 receptor. Clinical features of these patients resemble those seen in MC4R deficiency with, in addition, a variable phenotype of developmental delay with some autistic like features noted in some, but not all, patients.

Another summary of a recent book chapter by O´Rahilly S and Farooqi IS follows:

Mol Metab. 2015 Feb 7;4(4):287-98.

FTO is necessary for the induction of leptin resistance by high-fat feeding. Tung YC1, Gulati P1, Liu CH2, Rimmington D1, Dennis R1, Ma M1, Saudek V1, O’Rahilly S1, Coll AP1, Yeo GS1.1University of Cambridge Metabolic Research Laboratories and MRC Metabolic Diseases Unit, Wellcome-MRC Institute of Metabolic Science, Addenbrooke’s Hospital, Cambridge, UK.2Department of Physiology, Development and Neuroscience, University of Cambridge, Cambridge, UK.

Abstract

OBJECTIVE: Loss of function FTO mutations significantly impact body composition in humans and mice, with Fto-deficient mice reported to resist the development of obesity in response to a high-fat diet (HFD). We aimed to further explore the interactions between FTO and HFD and determine if FTO can influence the adverse metabolic consequence of HFD. METHODS: We studied mice deficient in FTO in two well validated models of leptin resistance (HFD feeding and central palmitate injection) to determine how Fto genotype may influence the action of leptin. Using transcriptomic analysis of hypothalamic tissue to identify relevant pathways affected by the loss of Fto, we combined data from co-immunoprecipitation, yeast 2-hybrid and luciferase reporter assays to identify mechanisms through which FTO can influence the development of leptin resistant states. RESULTS: Mice deficient in Fto significantly increased their fat mass in response to HFD. Fto (+/-) and Fto (-/-) mice remained sensitive to the anorexigenic effects of leptin, both after exposure to a HFD or after acute central application of palmitate. Genes encoding components of the NFкB signalling pathway were down-regulated in the hypothalami of Fto-deficient mice following a HFD. When this pathway was reactivated in Fto-deficient mice with a single low central dose of TNFα, the mice became less sensitive to the effect of leptin. We identified a transcriptional coactivator of NFкB, TRIP4, as a binding partner of FTO and a molecule that is required for TRIP4 dependent transactivation of NFкB. CONCLUSIONS: Our study demonstrates that, independent of body weight, Fto influences the metabolic outcomes of a HFD through alteration of hypothalamic NFкB signalling. This supports the notion that pharmacological modulation of FTO activity might have the potential for therapeutic benefit in improving leptin sensitivity, in a manner that is influenced by the nutritional environment. KEYWORDS: FTO, FaT mass and Obesity related; Fto; GWAS, Genome-wide association studies; HFD, high-fat diet; High-fat diet; Hypothalamus; ICV, intracerebroventricular injection; Irx3, Iroquois Homeobox 3; Leptin resistance; MEF, Mouse embryonic fibroblasts; NFкB; Ob-R, leptin receptor; PTPs, protein-tyrosine phosphatase; SNPs, single nucleotide polymorphisms; SOCS3; SOCS3, suppressor of cytokine signalling; TRIP4; Tlr4, Toll-like receptor 4; WAT, white adipose tissue; Y2H, Yeast two-hybrid

L3-1 Nutrient Sensing Nuclear Receptors Coordinate Liver Energy Balance
Session: L3-Friday Morning Plenary
Basic

David D Moore, PhD
Baylor College of Medicine, Houston, TX

Talk Description:

The liver has fundamentally different functions in the fed and fasted states. We have found that opposite, but complementary effects of the nutrient sensing nuclear receptors PPARalpha and FXR maintain liver energy balance over this cycle. In the fasted liver, PPARalpha activation transcriptionally induces autophagy, recycling cellular components to provide energy. PPARalpha activation also represses the energy expensive liver secretome. In contrast, FXR activation in the energy replete fed state represses autophagy, but supports secretion.

The summary of a recent paper by Dr. Moore et al. follows:

Nature. 2014 Dec 4;516(7529):112-5.

Nutrient-sensing nuclear receptors coordinate autophagy.

Lee JM1, Wagner M1, Xiao R1, Kim KH1, Feng D2, Lazar MA2, Moore DD1.1Department of Molecular and Cellular Biology, Baylor College of Medicine, Houston, Texas 77030, USA.2Division of Endocrinology, Diabetes, and Metabolism and the Institute for Diabetes, Obesity, and Metabolism, Perelman School of Medicine at the University of Pennsylvania, Philadelphia, Pennsylvania 19014, USA.

Abstract

Autophagy is an evolutionarily conserved catabolic process that recycles nutrients upon starvation and maintains cellular energy homeostasis. Its acute regulation by nutrient-sensing signalling pathways is well described, but its longer-term transcriptional regulation is not. The nuclear receptors peroxisome proliferator-activated receptor-α (PPARα) and farnesoid X receptor (FXR) are activated in the fasted and fed liver, respectively. Here we show that both PPARα and FXR regulate hepatic autophagy in mice. Pharmacological activation of PPARα reverses the normal suppression of autophagy in the fed state, inducing autophagic lipid degradation, or lipophagy. This response is lost in PPARα knockout (Ppara(-/-), also known as Nr1c1(-/-)) mice, which are partially defective in the induction of autophagy by fasting. Pharmacological activation of the bile acid receptor FXR strongly suppresses the induction of autophagy in the fasting state, and this response is absent in FXR knockout (Fxr(-/-), also known as Nr1h4(-/-)) mice, which show a partial defect in suppression of hepatic autophagy in the fed state. PPARα and FXR compete for binding to shared sites in autophagic gene promoters, with opposite transcriptional outputs. These results reveal complementary, interlocking mechanisms for regulation of autophagy by nutrient status.

 

L3-2 New Insights into Thyroid Hormone Signaling

Session: L3-Friday Morning Plenary
Basic

Anthony Neil Hollenberg, MD
Beth Israel Deaconess Med Center and Harvard Medical School, Boston, MA

Talk Description:

Thyroid Hormone exerts key effects on metabolism via its actions on diverse cell types. Central to interpreting its actions is the ability to understand how different cell types integrate circulating thyroid hormone levels. The use of novel mouse models and metabolite profiling has allowed for a new understanding of how different tissues respond to thyroid hormone to regulate metabolic function.

The summary of two recent papers by Dr. Hollenberg et al. follows:

Mol Cell Biol. 2014 May;34(9):1564-75.

Thyroid hormone signaling in vivo requires a balance between coactivators and corepressors. Vella KR1, Ramadoss P, Costa-E-Sousa RH, Astapova I, Ye FD, Holtz KA, Harris JC, Hollenberg AN.1Division of Endocrinology, Diabetes and Metabolism, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, Massachusetts, USA.

Abstract

Resistance to thyroid hormone (RTH), a human syndrome, is characterized by high thyroid hormone (TH) and thyroid-stimulating hormone (TSH) levels. Mice with mutations in the thyroid hormone receptor beta (TRβ) gene that cannot bind steroid receptor coactivator 1 (SRC-1) and Src-1(-/-) mice both have phenotypes similar to that of RTH. Conversely, mice expressing a mutant nuclear corepressor 1 (Ncor1) allele that cannot interact with TRβ, termed NCoRΔID, have low TH levels and normal TSH. We hypothesized that Src-1(-/-) mice have RTH due to unopposed corepressor action. To test this, we crossed NCoRΔID and Src-1(-/-) mice to create mice deficient for coregulator action in all cell types. Remarkably, NCoR(ΔID/ΔID) Src-1(-/-) mice have normal TH and TSH levels and are triiodothryonine (T(3)) sensitive at the level of the pituitary. Although absence of SRC-1 prevented T(3) activation of key hepatic gene targets, NCoR(ΔID/ΔID) Src-1(-/-) mice reacquired hepatic T(3) sensitivity. Using in vivo chromatin immunoprecipitation assays (ChIP) for the related coactivator SRC-2, we found enhanced SRC-2 recruitment to TR-binding regions of genes in NCoR(ΔID/ΔID) Src-1(-/-) mice, suggesting that SRC-2 is responsible for T(3) sensitivity in the absence of NCoR1 and SRC-1. Thus, T(3) targets require a critical balance between NCoR1 and SRC-1. Furthermore, replacement of NCoR1 with NCoRΔID corrects RTH in Src-1(-/-) mice through increased SRC-2 recruitment to T(3) target genes.

Mol Cell Biol. 2015 Feb;35(3):555-65.

NCoR1 and SMRT play unique roles in thyroid hormone action in vivo. Shimizu H1, Astapova I1, Ye F1, Bilban M2, Cohen RN3, Hollenberg AN4.1Division of Endocrinology, Diabetes and Metabolism, Beth Israel Deaconess Medical Center and Harvard Medical School, Boston Massachusetts, USA.2Department of Laboratory Medicine and Core Facility Genomics, Medical University of Vienna, Vienna, Austria.3Section of Endocrinology, University of Chicago, Chicago, Illinois, USA roncohen@medicine.bsd.uchicago.edu thollenb@bidmc.harvard.edu.4Division of Endocrinology, Diabetes and Metabolism, Beth Israel Deaconess Medical Center and Harvard Medical School, Boston Massachusetts, USA roncohen@medicine.bsd.uchicago.edu thollenb@bidmc.harvard.edu.

Abstract

NCoR1 (nuclear receptor corepressor) and SMRT (silencing mediator of retinoid and thyroid hormone receptors; NCoR2) are well-recognized coregulators of nuclear receptor (NR) action. However, their unique roles in the regulation of thyroid hormone (TH) signaling in specific cell types have not been determined. To accomplish this we generated mice that lacked function of either NCoR1, SMRT, or both in the liver only and additionally a global SMRT knockout model. Despite both corepressors being present in the liver, deletion of SMRT in either euthyroid or hypothyroid animals had little effect on TH signaling. In contrast, disruption of NCoR1 action confirmed that NCoR1 is the principal mediator of TH sensitivity in vivo. Similarly, global disruption of SMRT, unlike the global disruption of NCoR1, did not affect TH levels. While SMRT played little role in TH-regulated pathways, when disrupted in combination with NCoR1, it greatly accentuated the synthesis and storage of hepatic lipid. Taken together, these data demonstrate that corepressor specificity exists in vivo and that NCoR1 is the principal regulator of TH action. However, both corepressors collaborate to control hepatic lipid content, which likely reflects their cooperative activity in regulating the action of multiple NRs including the TH receptor (TR).

 

L4-1 Cancer, Bone and Muscle: It’s all about the Microenvironment

Session: L4-Friday Afternoon Plenary
Translational

Theresa Ann Guise, MD
Indiana University School of Medicine, Indianapolis, IN

Talk Description:

Bone metastases, common in advanced malignancy, cause significant morbidity: pain, fracture, and muscle weakness. Tumor cells in bone disrupt normal bone remodeling to fuel a feed-forward cycle of bone destruction. Tumors stimulate osteoclastic bone resorption, to release growth factors from the bone matrix, which act on tumor cells to fuel production of more osteolytic factors. These factors act systemically to induce oxidation and weakness in skeletal muscle by specific molecular mechanisms to be discussed.

The summary of a recent paper by Prof. Theresa Ann Guise follows:

J Bone Miner Res. 2015 Jan 15.

Single-Limb Irradiation Induces Local and Systemic Bone Loss in a Murine Model. Wright LE1, Buijs JT, Kim HS, Coats LE, Scheidler AM, John SK, She Y, Murthy S, Ma N, Sinex HJ, Bellido TM, Bateman TA, Mendonca MS, Mohammad KS, Guise TA.1Department of Medicine, Indiana University School of Medicine, Indianapolis, IN, USA.

Abstract

Increased fracture risk is commonly reported in cancer patients receiving radiotherapy, particularly at sites within the field of treatment. The direct and systemic effects of ionizing radiation on bone at a therapeutic dose are not well characterized in clinically relevant animal models. Using twenty-week male C57Bl/6 mice, effects of irradiation (right hindlimb; 2 Gy) on bone volume and microarchitecture were evaluated prospectively by microcomputed tomography and histomorphometry and compared to contralateral-shielded bone (left hindlimb) and non-irradiated control bone. One-week post-irradiation, trabecular bone volume declined in irradiated tibiae (-22%; p < 0.0001) and femora (-14%; p = 0.0586) and microarchitectural parameters were compromised. Trabecular bone volume declined in contralateral tibiae (-17%; p = 0.003), and no loss was detected at the femur. Osteoclast number, apoptotic osteocyte number and marrow adiposity were increased in irradiated bone relative to contralateral and non-irradiated bone, while osteoblast number was unchanged. Despite no change in osteoblast number one-week post-irradiation, dynamic bone formation indices revealed a reduction in mineralized bone surface and a concomitant increase in unmineralized osteoid surface area in irradiated bone relative to contralateral and non-irradiated control bone. Further, dose- and time-dependent calvarial culture and in vitro assays confirmed that calvarial osteoblasts and osteoblast-like MC3T3 cells were relatively radioresistant, while calvarial osteocyte and osteocyte-like MLO-Y4 cell apoptosis was induced as early as 48h post-irradiation (4 Gy). In osteoclastogenesis assays, radiation exposure (8 Gy) stimulated murine macrophage RAW264.7 cell differentiation and co-culture of irradiated RAW264.7 cells with MLO-Y4 or murine bone marrow cells enhanced this effect. These studies highlight the multi-faceted nature of radiation-induced bone loss by demonstrating direct and systemic effects on bone and its many cell types using clinically relevant doses and have important implications for bone health in patients treated with radiation therapy. L4-2 Defining the Genetic Basis of Benign and Malignant Adrenocortical Tumors

Program: Plenary
Session: L4-Friday Afternoon Plenary
Translational

Jerome Yves Bertherat, MD, PhD
INSERM U 1016, CNRS 8104, Institut Cochin, Paris Descartes University, Paris, France

Talk Description: There is a variety of adrenocortical tumors that can be responsible for different cortisol excess levels. Genomics allowed recently many progress in this field. Bilateral tumors (macronodular and micronodular adrenal hyperplasia) are often caused by germline alterations (ARMC5, PRKAR1A). Unilateral tumors (adenoma or cancer) are caused by somatic alterations (CTNNB1, TP53, ZNRF3, MEN1, RB1, PRKACA, …). The importance of these advances for early diagnosis, tumors classification improvement, and development of new therapies will be discussed.

The summary of a recent paper by Prof. Jerome Yves Bertherat follows:

The Genetics of Adrenocortical Tumors. Espiard S1, Bertherat J2.

Endocrinol Metab Clin North Am. 2015 Jun;44(2):311-334. doi: 10.1016/j.ecl.2015.02.004. 1Cochin Institut, INSERM U1016, 24 rue du Faubourg Saint Jacques, Paris 75014, France; Cochin Institut, CNRS UMR8104, 24 rue du Faubourg Saint-Jacques, Paris 75014, France; Paris Descartes University, 12 rue de l’Ecole de Médecine, Paris 75006, France.2Cochin Institut, INSERM U1016, 24 rue du Faubourg Saint Jacques, Paris 75014, France; Cochin Institut, CNRS UMR8104, 24 rue du Faubourg Saint-Jacques, Paris 75014, France; Paris Descartes University, 12 rue de l’Ecole de Médecine, Paris 75006, France; Endocrinology Department, Center for Rare Adrenal Diseases, Hôpital Cochin, Assistance Publique Hôpitaux de Paris, 27 Rue du Fg-St-Jacques, Paris F-75014, France. Electronic address:

Abstract

Advances in genomics accelerated greatly progress in the study of the genetics adrenocortical tumors. Bilateral nodular hyperplasias causing Cushing’s syndrome are frequently caused by germline alterations leading to cAMP/PKA pathway activation (micronodular) and ARMC5 (chromosome location 16p11.2) inactivation (macronodular). Somatic mutations of β-catenin and PRKACA (protein kinase, cAMP-dependent, catalytic, alpha) are observed in non secreting or cortisol producing adenomas, respectively. Alterations of the β-catenin (CTNN1B, ZNFR3) or TP53 pathways are found in carcinomas. Mutations in cancers are more common in aggressive tumors and correlate with transcriptome or methylation profiles. Identification of these alterations helps to refine the molecular classification of these tumors and to develop molecular diagnostic tools.

 

L5-1 microRNAs: Key Regulators of Cancer Initiation and Progression

Session: L5-Saturday Morning Plenary
Translational

Carlo M Croce, M.D
Ohio State University, Columbus, OH

Talk Description: Our understanding of non-coding (nc)-RNAs expression patterns and function in normal or neoplastic human cells is now emerging. nc-RNAs expression profiles of tumors have been shown to be potential tools for cancer prognosis. Furthermore, recent data are consistent with the hypothesis that ncRNAs play a substantial and unanticipated role in the pathogenesis of human cancers.

Methods Mol Biol. 2015;1297:137-52. Functional assays for specific targeting and delivery of RNA nanoparticles to brain tumor. Lee TJ1, Haque F, Vieweger M, Yoo JY, Kaur B, Guo P, Croce CM.1Department of Molecular Virology, Immunology and Medical Genetics, Comprehensive Cancer Center, The Ohio State University, 1080 Biomedical Research Tower, 460 W. 12th Ave., Columbus, OH, 43210, USA.

Abstract

Cumulative progress in nanoparticle development has opened a new era of targeted delivery of therapeutics to cancer cells and tissue. However, developing proper detection methods has lagged behind resulting in the lack of precise evaluation and monitoring of the systemically administered nanoparticles. RNA nanoparticles derived from the bacteriophage phi29 DNA packaging motor pRNA have emerged as a new generation of drugs for cancer therapy. Multifunctional RNA nanoparticles can be fabricated by bottom-up self-assembly of engineered RNA fragments harboring targeting (RNA aptamer or chemical ligand), therapeutic (siRNA, miRNA, ribozymes, and small molecule drugs), and imaging (fluorophore, radiolabels) modules. We have recently demonstrated that RNA nanoparticles can reach and target intracranial brain tumors in mice upon systemic injection with little or no accumulation in adjacent healthy brain tissues or in major healthy internal organs. Herein, we describe various functional imaging methods (fluorescence confocal microscopy, flow cytometry, fluorescence whole body imaging, and magnetic resonance imaging) to evaluate and monitor RNA nanoparticle targeting to intracranial brain tumors in mice. Such imaging techniques will allow in-depth evaluation of specifically delivered RNA therapeutics to brain tumors.

J Hematol Oncol. 2015 Feb 20;8(1):12.

Role of microRNA in chronic lymphocytic leukemia onset and progression. Balatti V1, Pekarky Y2, Croce CM3.1Department of Molecular Virology, Immunology and Medical Genetics, Comprehensive Cancer Center, Wexner Medical Center, The Ohio State University, Columbus, OH, USA. Veronica.Balatti@osumc.edu.2Department of Molecular Virology, Immunology and Medical Genetics, Comprehensive Cancer Center, Wexner Medical Center, The Ohio State University, Columbus, OH, USA. Yuri.Pekarsky@osumc.edu.3Department of Molecular Virology, Immunology and Medical Genetics, Comprehensive Cancer Center, Wexner Medical Center, The Ohio State University, Columbus, OH, USA. Carlo.Croce@osumc.edu.

Abstract

B-cell chronic lymphocytic leukemia (CLL) is the most common human leukemia occurring as indolent or aggressive form. CLL clinical features and genetic abnormalities are well documented, but molecular details are still under investigation. MicroRNAs are small non-coding RNAs involved in several cellular processes and expressed in a tissue-specific manner. MicroRNAs regulate gene expression, and their deregulation can alter expression levels of genes involved in development/progression of tumors. In CLL, microRNAs can function as oncogenes or tumor suppressors and can also serve as markers for CLL onset/progression. Here, we discuss the most recent findings about the role of microRNAs in CLL and how this knowledge can be used to identify new biomarkers and treatment approaches.

L5-2 The Wiring Diagram for Hunger: Using Neuron-Specific Tools to Discover its Basis
Session: L5-Saturday Morning Plenary
Basic
Bradford Barr Lowell, MD, PHD
Beth Israel Deaconess Med Ctr, Boston, MA

Talk Description:

Efforts to elucidate neural circuits regulating hunger have been stymied by the complexity of brain interconnections. With recently developed circuit-specific approaches, this complexity can now be “solved”. We are using optogenetics, chemogenetics and monosynaptic rabies mapping, rendered cell-specific by lines of mice expressing cre-recombinase, to map synaptic connections between neurons and to probe function of identified neural circuits. The ultimate goal is to uncover the neurobiological basis, i.e. the wiring diagram, for hunger and satiety.

Curr Biol. 2014 Dec 1;24(23):R1111-6. The hypothalamus. Saper CB1, Lowell BB2.1Department of Neurology, Beth Israel Deaconess Medical Center and Harvard Medical School, Boston MA 02215, USA; Program in Neuroscience, Beth Israel Deaconess Medical Center and Harvard Medical School, Boston MA 02215, USA. Electronic address: csaper@bidmc.harvard.edu.2Division of Endocrinology, Beth Israel Deaconess Medical Center and Harvard Medical School, Boston MA 02215, USA; Program in Neuroscience, Beth Israel Deaconess Medical Center and Harvard Medical School, Boston MA 02215, USA.

Abstract

The hypothalamus is one of the oldest and smallest parts of the brain, constituting just 4 gm of the 1400 gm of adult human brain weight. And yet this tiny area contains highly conserved neural circuitry that controls basic life functions: these include energy metabolism, from feeding through digestion, metabolic control, and energy expenditure; fluid and electrolyte balance, from drinking through fluid absorption and excretion; thermoregulation, from choice of environment through heat production and conservation, and fever responses; wake-sleep cycles and emergency responses to stressors in the environment; and reproduction, from reproductive hormone control through mating, pregnancy, birth, and suckling. In this Primer, we will give an overview of the structure of the hypothalamus, and outline what we know about how that relates to its functional circuitry.

L6-1 New Players in the Regulation of Human Puberty

Session: L6-Saturday Afternoon Plenary
Translational

Ana Claudia Latronico, MD, PhD
Sao Paulo University, Sao Paulo, Brazil

Talk Description: A pivotal event in the onset of puberty in humans is the re-emergence of pulsatile release of gonadotropin-releasing hormone (GnRH) from the hypothalamic neurons. Pathways governing GnRH ontogeny and physiology have been discovered by studying animal models and humans with reproductive disorders. Recent human studies implicated the activation of the kisspeptin and its cognate receptor (KISS1- KISS1R), and the inactivation of the makoring ring finger 3 (MKRN3) in the premature reactivation of the GnRH secretion, causing central precocious puberty. In addition, epigenetic repression of key inibitory loci may play a fundamental role in the initiation of puberty.

The summary of a recent paper by Dr. Latronico et al. follows

J Clin Endocrinol Metab. 2014 Jun;99(6):E1097-103. doi: 10.1210/jc.2013-3126. Central precocious puberty that appears to be sporadic caused by paternally inherited mutations in the imprinted gene makorin ring finger 3.Macedo DB1, Abreu AP, Reis AC, Montenegro LR, Dauber A, Beneduzzi D, Cukier P, Silveira LF, Teles MG, Carroll RS, Junior GG, Filho GG, Gucev Z, Arnhold IJ, de Castro M, Moreira AC, Martinelli CE Jr, Hirschhorn JN, Mendonca BB, Brito VN, Antonini SR, Kaiser UB, Latronico AC.1Unidade de Endocrinologia do Desenvolvimento e Laboratório de Hormônios e Genética Molecular/LIM42, Disciplina de Endocrinologia (D.B.M., A.P.A., L.R.M., D.B., P.C., L.F.G.S., M.G.T., I.J.P.A., B.B.M., V.N.B., A.C.L.), Hospital das Clínicas da Faculdade de Medicina da Universidade de São Paulo, São Paulo, SP, Brasil, 05403-900; Departamento de Puericultura e Pediatria (A.C.S.R., M.C., A.C.M., C.E.M., S.R.A.), Faculdade de Medicina de Ribeirão Preto, da Universidade de São Paulo, Ribeirão Preto, SP, Brasil, 14049900; Division of Endocrinology, Diabetes, and Hypertension (A.P.A., R.S.C., U.B.K.), Brigham and Women’s Hospital and Harvard Medical School and Division of Endocrinology, (A.D., J.N.H.) Boston Children’s Hospital, Boston, Massachusetts 02115, Program in Medical and Population Genetics Broad Institute (A.D., J.N.H.), Cambridge, Massachusetts 02142; Unidade de Endocrinologia Pediátrica (G.G.J., G.G.F.), Universidade de Campinas, SP, Brasil, 13084-970; and Medical Faculty Skopje (Z.G.), 50 Divizija BB, 1000 Skopje, Macedonia.

Abstract

CONTEXT: Loss-of-function mutations in makorin ring finger 3 (MKRN3), an imprinted gene located on the long arm of chromosome 15, have been recognized recently as a cause of familial central precocious puberty (CPP) in humans. MKRN3 has a potential inhibitory effect on GnRH secretion.OBJECTIVES: The objective of the study was to investigate potential MKRN3 sequence variations as well as copy number and methylation abnormalities of the 15q11 locus in patients with apparently sporadic CPP.SETTING AND PARTICIPANTS: We studied 215 unrelated children (207 girls and eight boys) from three university medical centers with a diagnosis of CPP. All but two of these patients (213 cases) reported no family history of premature sexual development. First-degree relatives of patients with identified MKRN3 variants were included for genetic analysis.MAIN OUTCOME MEASURES: All 215 CPP patients were screened for MKRN3 mutations by automatic sequencing. Multiplex ligation-dependent probe amplification was performed in a partially overlapping cohort of 52 patients.RESULTS: We identified five novel heterozygous mutations in MKRN3 in eight unrelated girls with CPP. Four were frame shift mutations predicted to encode truncated proteins and one was a missense mutation, which was suggested to be deleterious by in silico analysis. All patients with MKRN3 mutations had classical features of CPP with a median age of onset at 6 years. Copy number and methylation abnormalities at the 15q11 locus were not detected in the patients tested for these abnormalities. Segregation analysis was possible in five of the eight girls with MKRN3 mutations; in all cases, the mutation was inherited on the paternal allele.CONCLUSIONS: We have identified novel inherited MKRN3 defects in children with apparently sporadic CPP, supporting a fundamental role of this peptide in the suppression of the reproductive axis.

L6-2 PCOS: New Insights into Molecular Mechanisms

Session: L6-Saturday Afternoon Plenary
Translational

Jerome F Strauss III, MD, Ph.D.
Virginia Commonwealth University, Richmond, VA

Talk Description: Recent discoveries in genetics and functional genomics relevant to PCOS will be reviewed, and their implications for the pathophysiology, diagnosis and treatment of PCOS will be discussed.

The summary of a recent paper by Dr. Jerome F Strauss III follows:

Trends Endocrinol Metab. 2015 Mar;26(3):118-24. McAllister JM1, Legro RS2, Modi BP3, Strauss JF 3rd3. Functional genomics of PCOS: from GWAS to molecular mechanisms.1Department of Pathology, Penn State Hershey College of Medicine, Hershey, PA, USA; Department of Obstetrics and Gynecology, Penn State Hershey College of Medicine, Hershey, PA, USA. Electronic address: jmcallister@psu.edu.2Department of Obstetrics and Gynecology, Penn State Hershey College of Medicine, Hershey, PA, USA.3Department of Obstetrics and Gynecology, Virginia Commonwealth University, Richmond, VA, USA.

Abstract

Polycystic ovary syndrome (PCOS) is a common endocrinopathy characterized by increased ovarian androgen biosynthesis, anovulation, and infertility. PCOS has a strong heritable component based on familial clustering and twin studies. Genome-wide association studies (GWAS) identified several PCOS candidate loci including LHCGR, FSHR, ZNF217, YAP1, INSR, RAB5B, and C9orf3. We review the functional roles of strong PCOS candidate loci focusing on FSHR, LHCGR, INSR, and DENND1A. We propose that these candidates comprise a hierarchical signaling network by which DENND1A, LHCGR, INSR, RAB5B, adapter proteins, and associated downstream signaling cascades converge to regulate theca cell androgen biosynthesis. Future elucidation of the functional gene networks predicted by the PCOS GWAS will result in new diagnostic and therapeutic approaches for women with PCOS.

Comments

Hyperandrogenemia and PCOS. PCOS is a common disorder that is reported to affect 5–7% women of reproductive age. The hyperandro-genemia is widely believed to be primarily of ovarian theca cell origin, although the adrenal zona reticularis contributes androgens, mainly dehydroepiandrosterone (DHEA), in approximately 25% of cases. The interconnectedness between the two androgen-secreting organs in PCOS is supported by the suppression of both adrenal and ovarian androgens by oral contraception or insulin-sensitizing agents. This review focuses on the genetic component underlying androgen excess in PCOS.

The phenotype of human theca cells from normal and PCOS ovaries. PCOS theca secretes greater amounts of androgen than theca tissue or cells from regularly ovulating women via increased expression of the rate-limiting enzyme in androgen biosynthesis, P450 17a-hydroxylase/17,20 lyase, encoded by the CYP17A1 gene. An array of structural proteins, cell cycle proteins, and transcription factors, including the transcription factor GATA-6, show increased expression. GATA-6 was subsequently shown to be a transcriptional activator of the main P450 steroidogenic enzymes involved in androgen biosynthesis, including CYP17A1 and CYP11A1.

The genetics of PCOS. PCOS is a heterogeneous disorder with strong evidence for a genetic component. They reviewed the functional roles of strong PCOS candidate loci identified in the GWAS studies, and subsequently replicated in other populations, focusing on FHSR, LHCGR, INSR, and DENND1A.

Coding sequence variants in FSHR, LHCGR, and INSR associated with PCOS and their functional significance. The FSH receptor (FSHR). Inactivating mutations in the FSHR gene lead to hypergonadotropic hypogonadism, and in the absence of a functional FSHR follicular development is generally halted at the preantral stage. Conversely, mutations in the transmembrane helices and the extracellular domain of FSHR are associated with spontaneous ovarian hyperstimulation síndrome. There is no clear explanation for the correlation between genotype of the FSHR variants and FSH levels or response to gonadotropins, and, by extension, how exactly these variants contribute to PCOS is not clear.

The LH/CG receptor (LHCGR). In the ovary, induction of the LHCGR during granulosa cell differentiation is necessary for the pre-ovulatory follicle to respond to the mid-cycle LH surge resulting in ovulation. The fact that known activating mutations of the LHCGR that cause precocious puberty in males because of premature Leydig cell activa-tion and testosterone production are not associated with hyperandrogenemia of ovarian origin in women calls into question the notion that ‘hyper-responsive’ LHCGR isoforms are related to theca cell dysfunction.

The insulin receptor (INSR). The importance of insulin signaling in PCOS is evident through the HAIRAN syndrome (hyperandrogenism, insulin resistance acanthosis nigricans), which is a sub-phenotype of PCOS characterized by severe insulin resistance.

Collectively, candidate gene association studies provide some additional support for the notion that the FSHR, LHCGR, and INSR loci are associated with PCOS, consistent with the GWAS and replication studies noted above. However, there is so far no convincing evidence that the variants identified in these genes are functionally significant and contribute to the PCOS theca cell phenotype.

DENND1A. This gene is a novel starting point for dissecting the molecular mechanisms underlying PCOS. DENND1A encodes the plasma-membrane protein connecdenn 1, which is involved in clathrin binding, endocytic processes, and receptor recycling. DENND1A functions as a guanine nucleotide exchange factor that interacts with members of the Rab family of small GTPases. DENND1A is thought to be involved in clathrin-mediated endocytosis, facilitating internalization of proteins and lipids, receptor recycling, and membrane trafficking. DENND1A has also been associated with phosphatidylinositol-3-phosphate and with endocytosis/endosome proteins The DENND1A gene yields two principal transcripts via alternative splicing: DENND1A variant 1 (DENND1A.V1), which encodes a 1009 amino acid protein with a large C-terminal proline-rich domain, and DENND1A.V2 that encodes a 559 AA protein that lacks the proline-rich domain, and includes a distinct C-terminal 33 AA sequence that differs from DENND1A.V1. DENND1A is highly expressed in ovarian theca cells and the adrenal zona reticularis, both androgen-producing tissues. Until recently little was known about DENND1A expression in cells and tissues related to reproduction. DENND1A.V2 in PCOS theca cells. DENND1A.V2 expression is increased in PCOS theca cells. Data are available providing strong evidence that increased DENND1A.V2 expression mediates augmented CYP17A1 gene expression and androgen biosynthesis in PCOS theca cells. DENND1A.V2 acts in concert with genetic variants of FSHR, LHCGR, and INSR to modulate signal transduction pathways that promote theca androgen synthesis. It would be a novel therapeutic and diagnostic target for PCOS.

Exosomes are small nucleic acid rich vesicles shed into blood and urine, which provide a source of RNA that can be conveniently utilized for diagnostic implications. Studies comparing DENNDD1.V2 mRNA in urine exosomes isolated from normal cycling and PCOS women demonstrated that DENND1A.V2 RNA is increased in PCOS compared to normally cycling women. These experiments provided the first evidence to suggest that increased urine exosomal DENND1A.V2 could be diagnostic for PCOS. A diagnostic method based on urine exosomal RNA would have value in settings when non-invasive diagnostics are needed, such as in prepubertal and adolescent females. However, this test requires extensive validation to assess its sensitivity, specificity, and positive and negative predictive value. Possible mechanisms for overexpression include copy-number variation (CNV), increased promoter activity, the influence of microRNAs on protein expression, or splicing variation favoring the production of DENND1A.V2. PCOS is associated with excess adrenal androgen levels in approximately 25% of women, consistent with an adrenal steroidogenic abnormality. The localization of DENND1A.V2 in the human adrenal zona reticularis provides evidence that alterations in the expres- sion of this isoform may contribute to the adrenal steroido- genic abnormality in addition to the theca derangement in androgen production. DENND1A SNPs have been reported to be associated with diverse phenotypes found in PCOS women, including elevated insulin levels and endometrioid carcinoma. In addition, quantitative trait analyses have linked the DENND1A locus with glucose and insulin levels and body weight in humans. Thus, DENND1A.V2 could possibly contribute directly to or modify several PCOS phenotypes, which heretofore have not been tied to a single genetic locus. Therefore, an additional area of further exploration is the role of DENND1A variants in predicting response to common treatments for women with PCOS. Hyperandrogenism is clearly associated with the development of hirsutism in women with PCOS, and furthermore, is an important predictor of anovulation and failure to respond to ovulation induction. Thus, DENNDIA may be a particularly useful molecular marker for pharmacogenomics studies. Recent large multicenter studies of ovulation induction methods in women with PCOS and anovulatory infertility will certainly aid these studies.

L7-1 Enteroendocrine Control of Metabolism-Physiological Insights and Therapeutic Implications

Session: L7-Sunday Morning Plenary
Clinical

Daniel J Drucker, MD
Mt Sinai Hospital, Toronto, ON, Canada

Talk Description: The enteroendocrine system represents the largest endocrine network that controls hormonal signals regulating ingestion, digestion, absorption and disposal of nutrients. The mechanisms through which these hormones exert their actions will be discussed. We also demonstrate the benefits and potential risks of gut-based therapies for the treatment of obesity, diabetes and intestinal disorders.

The summary of a recent paper by Dr. Daniel J Drucker III follows:

Diabetes. 2015 Feb;64(2):317-26.

Drucker DJ. Deciphering metabolic messages from the gut drives therapeutic innovation: the 2014 Banting Lecture. Department of Medicine, Mount Sinai Hospital and the Lunenfeld-Tanenbaum Research Institute, University of Toronto, Toronto, Ontario, Canada

Abstract

The Banting Medal for Scientific Achievement is the highest scientific award of the American Diabetes Association (ADA). Given in memory of Sir Frederick Banting, one of the key investigators in the discovery of insulin, the Banting Medal is awarded annually for scientific excellence, recognizing significant long-term contributions to the understanding, treatment, or prevention of diabetes. Daniel J. Drucker, MD, of the Department of Medicine, Mount Sinai Hospital and the Lunenfeld-Tanenbaum Research Institute in Toronto, Ontario, Canada, received the prestigious award at the ADA’s 74th Scientific Sessions, 13-17 June 2014, in San Francisco, California. He presented the Banting Lecture, “Deciphering Metabolic Messages From the Gut Drives Therapeutic Innovation,” on Sunday, 15 June 2014. Gut peptides convey nutrient-regulated signals to the enteric nervous system and to distal organs, acting as circulating hormones secreted in the basal and postprandial state. Here an overview of the actions of glucagon-like peptide (GLP)-1 and GLP-2, the two major enteroendocrine L-cell peptides are provided. The endogenous physiological actions of GLP-1 have been delineated using antagonists and Glp1r(-/-) mice and include the control of islet hormone secretion in a glucose-dependent manner, leading to improvement of fasting and postprandial glucose homeostasis. GLP-1 receptors (GLP-1Rs) are also widely distributed in multiple extrapancreatic organs, providing a mechanistic explanation for the nonglycemic actions attributed to GLP-1. The multiple metabolic actions of GLP-1 enable reduction of glycemia and body weight in diabetic and obese subjects, providing the opportunity to reduce glycemia in human subjects with diabetes with a low risk of hypoglycemia. GLP-2 plays a key role in the control of energy absorption and in the integrity of the intestinal mucosa, and a GLP-2R agonist, teduglutide, is now used for augmentation of energy absorption in parenteral nutrition-dependent subjects with short bowel syndrome. GLP-1 and GLP-2 are both cleaved by dipeptidyl peptidase-4 (DPP-4); hence, inhibition of DPP-4 activity enables yet another pathway for potentiation of incretin action and the therapy for type 2 diabetes. Here, a 30-year experience on the elucidation of gut hormone action is reviewed and, wherever possible, highlight therapeutic implications of preclinical studies and future opportunities for incretin research is commented.

Comments

THE PROGLUCAGON-DERIVED PEPTIDES. The glucagon-like peptide (GLP)-1 and GLP-2 are the two major enteroendocrine products produced by L-cell in the gut.

THE ACTIONS OF GLP-1. GLP-1(7-37) potently increases cAMP formation, insulin secretion, and insulin gene expression in

β-cells. The first GLP-1 receptor (GLP-1R) agonist, exenatide, was approved for clinical use in April 2005, followed by approval of the first

dipeptidyl peptidase-4 (DPP-4) inhibitor, sitagliptin, in October 2006.

However, they do not improve the unexpected actions of metformin, which not only inhibits hepatic glucose production and stimulates GLP-1 secretion but in murine studies also potentiates incretin action at the β-cell.

PHYSIOLOGY OF GLP-1 ACTION: AN INITIAL AND SUSTAINED FOCUS ON β-CELLS. Although the GLP-1R antagonist exendin had been identified, it is a weak partial agonist and inverse agonist. Studies have demonstrated that basal GLP-1R signaling is essential for control of glucagon and insulin secretion in rodents and humans. Indeed, the initial concepts of GLP-1 as a postprandial regulator of insulin secretion have now been extended to encompass a role for basal GLP-1 in the control of glycemia, even in the interprandial or fasting state.

The relative importance of the β-cell versus the α-cell versus the central nervous system (CNS) as a target for the glucoregulatory actions of endogenous GLP-1 continues to be a subject of debate. despite the importance of neural GLP-1R signaling networks for aspects of GLP-1 action relevant to control of appetite and body weight, the β-cell, and not the brain, is essential for GLP-1R–dependent glucoregulation.

GLP-1 CONTROLS β-CELL PROLIFERATION AND SURVIVAL. The pharmacological and physiological importance of GLP-1R signaling for cell survival has been established, a paradigm replicated in studies of GLP-1R+ neurons in the CNS. GLP-1R agonists uniquely augment β-cell function, simultaneously increasing the expression of molecules required for differentiated b-cell function and enhancing insulin biosynthesis and secretion while triggering pathways ensuring the survival and/or proliferation of stressed β-cells.

DPP-4. Mechanisms through which DPP-4i lower blood glucose.

DPP-4i prevent the degradation of GLP-1 and GIP, thereby augmenting signaling through their cognate receptors (GLP-1R and GIPR), resulting in stimulation of insulin and inhibition of glucagon secretion. It was demonstrated that metformin acutely increased GLP-1 secretion and acted as an incretin sensitizer, both directly in islet cells in vitro and in mice. Metformin increased the expression of the GLP-1Rs and GIPRs in mouse islets, actions requiring the transcription factor peroxisome proliferator–activated receptor-a. Combining DPP-4 inhibitors with metformin for the treatment of diabetes represents a logical approach to therapy through unexpected mechanistic interactions.

GLP-2: A NOVEL THERAPY FOR SHORT BOWEL SYNDROME. Proglucagon-derived peptides exhibited bowel growth factor activity. A GLP-2R agonist, teduglutide, was approved for the treatment of short bowel syndrome.

NONGLYCEMIC ACTIONS OF GLP-1: FOCUS ON THE CARDIOVASCULAR SYSTEM- Native GLP-1 and GLP-1(9-36) both improved ventricular function in ischemic ventricles ex vivo, through non–GLP-1R–dependent mechanisms of action, likely through regulation of blood flow.

THE FUTURE OF INCRETIN-BASED SCIENCE

The first 30 years of incretin-based science have witnessed an explosive growth in our understanding of how GLP-1 controls glucose homeostasis and body weight. Nevertheless, the widespread distribution of extrapancreatic GLP-1Rs has been accompanied by delineation of multiple nonglycemic actions of incretin hormones, many of which may have considerable clinical relevance. The therapeutic potential of GLP-1–based therapies may now be extended to the treatment of obesity, raising new scientific questions relevant to mechanisms of action in the CNS and introducing additional questions about long-term safety in a nondiabetic population.

 

7-2 Antihyperglycemic Therapy and Cardiovascular Disease: In Search of the Holy Grail

Session: L7-Sunday Morning Plenary
Basic

Silvio E Inzucchi, MD
Yale School of Medicine, New Haven, CT

Talk Description: The link between Type 2 diabetes and cardiovascular disease will be reviewed and placed into the context of the multiplicity of risk factors often encountered in this disease. The impact of glucose lowering therapy on cardiovascular events will next be described, both as regards to HbA1c targets and specific therapeutic classes. Ongoing clinical trials examining this important clinical question will be also discussed.

 

L8-1 Mechanisms of Androgen Dependence in Progressive Prostate Cancer

Session: L8-Sunday Afternoon Plenary
Basic

Karen E Knudsen, PhD
Thomas Jefferson University, Philadelphia, PA

Talk Description: There is increasing evidence that hormone action in cancer cells modulates the response to DNA damage. Conversely, a number of DNA repair factors have been found at sites of hormone receptor activity, suggesting that interplay between the DNA repair and hormone signaling pathways may have significant effects on the response to genomic insult and cancer cell survival. These mechanisms, and the therapeutic impact of hormone-DNA repair interplay, will be discussed.

The summary of a recent paper by Prof. Karen E Knudsen III follows:

Cancer Discov. 2014 Oct;4(10):1126-39.

Beyond DNA repair: DNA-PK function in cancer. Goodwin JF, Knudsen KE. Department of Cancer Biology, Department of Urology, Department of Radiation Oncology, Kimmel Cancer Center, Thomas Jefferson University, Philadelphia, Pennsylvania.

Abstract.

The DNA-dependent protein kinase (DNA-PK) is a pivotal component of the DNA repair machinery that governs the response to DNA damage, serving to maintain genome integrity. However, the DNA-PK kinase component was initially isolated with transcriptional complexes, and recent findings have illuminated the impact of DNA-PK-mediated transcriptional regulation on tumor progression and therapeutic response. DNA-PK expression has also been correlated with poor outcome in selected tumor types, further underscoring the importance of understanding its role in disease. Herein, the molecular and cellular consequences of DNA-PK are considered, with an eye toward discerning the rationale for therapeutic targeting of DNA-PK. SIGNIFICANCE: Although DNA-PK is classically considered a component of damage response, recent findings illuminate damage-independent functions of DNA-PK that affect multiple tumor-associated pathways and provide a rationale for the development of novel therapeutic strategies.

Comments

Identification of defined roles for DNA-PK in tumor associated processes including genomic stability, hypoxia, metabolism, inflammatory response, and transcription supports targeting DNA-PK as a therapeutic intervention in human malignancy. The therapeutic efficacy of targeting DNA-PK has classically been gauged through potential to prevent repair of damage. DNA-PK regulates repair of double strand breaks through the non-homologous end joining (NHEJ) DNA double strand break repair pathway. Consistent with this concept, synthetic lethal interactions have been identified between DNA-PKcs and multiple damage response factors including ATM, and numerous components of homologous recombination (HR)-mediated repair, suggesting that DNA-PK inhibitors may show efficacy as targeted agents for treatment of damage repair-defective human cancers. While many of the original DNA-PK inhibitors suffered from lack of specificity and poor pharmacokinetics, more recently developed inhibitors, including NU7026 and NU7441, demonstrate improved specificity and pharmaceutical properties and have been used preclinically to explore the impact of DNA-PK inhibition in multiple tumor types. However, recent evidence implicating DNA-PK in pathways outside of NHEJ may help define novel strategies of therapeutic intervention, a concept supported by the reality that all of the agents targeting DNA-PK currently in clinical trial reflect the dynamic ability of DNA-PK to impinge on multiple pathways. CC-122 is a DNA-PK inhibitor in phase I clinical trial (NCT01421524) for solid tumors, Non-Hodgkin’s lymphoma, and multiple myeloma. CC-122 is described as a pleiotropic pat hway modifier, echoing the ability of DNA-PK to modulate numerous pathways outside of double strand break repair. The two additional DNA-PK targeting molecules currently in clinical trial target both DNA-PK and the PI3K/Akt/mTOR signaling pathway. CC-115 is a dual inhibitor of DNA-PK and mTOR, and is currently in phase I trial (NCT01353625) for advanced solid tumors and hematologic malignancies while ZSTK474 is a PI3K inhibitor that also inhibits DNA-PK currently in phase I trials (NCT01280487 and NCT01682473) for advanced solid malignancies. As the only DNA-PK-targeting compounds currently in clinical trial impinge on pathways outside of DNA-PK-mediated damage repair, it will be important to consider novel DNA-PK regulated pathways when designing therapeutic strategies. For example, as a positive feedback circuit involving AR and DNA-PK drives PCa progression and survival, next generation means of targeting AR activity in combination with a DNA-PK inhibitor may represent a novel node of therapeutic intervention in advanced PCa. Though directly targeting cancer tissue is always important to consider with double strand break-inducing agents to avoid reduction of the therapeutic ratio, such combination strategies could afford new opportunities for precision therapies since DNA-PK seems to impact multiple tumor types.

L8-2 Guiding Hormonal Signals through Anchored Enzyme Complexes

Session: L8-Sunday Afternoon Plenary
Basic

John D Scott, PHD
Howard Hughes Med Inst, Seattle, WA

Talk Description: Dr. Scott’s talk will explore the spatial and temporal resolution of cell signaling events. Using a combined genetic, molecular and cell biology approach he and his colleagues have uncovered some of the mechanisms that cells use to constrain hormone induced cellular responses. These signals proceed through a family of signal organizing elements called AKAPs.

The summary of a recent paper by Prof. John D Scott follows:

Methods Mol Biol. 2015;1294:137-50.

Selective disruption of the AKAP signaling complexes. Kennedy EJ, Scott JD. Department of Pharmaceutical and Biomedical Sciences, University of Georgia College of Pharmacy, Athens, GA, USA.

Abstract

Synthesis of the second messenger cAMP activates a variety of signaling pathways critical for all facets of intracellular regulation. Protein kinase A (PKA) is the major cAMP-responsive effector. Where and when this enzyme is activated has profound implications on the cellular role of PKA. A-Kinase Anchoring Proteins (AKAPs) play a critical role in this process by orchestrating spatial and temporal aspects of PKA action. A popular means of evaluating the impact of these anchored signaling events is to biochemically interfere with the PKA-AKAP interface. Hence, peptide disruptors of PKA anchoring are valuable tools in the investigation of local PKA action. This article outlines the development of PKA isoform-selective disruptor peptides, documents the optimization of cell-soluble peptide derivatives, and introduces alternative cell-based approaches that interrogate other aspects of the PKA-AKAP interface.

The summary of a second recent paper by Prof. John D Scott follows:

J Biol Chem. 2015 Apr 16.

A-Kinase Anchoring Protein 79/150 recruits Protein Kinase C to phosphorylate Roundabout receptors. Samelson BK1, Gore BB2, Whiting JL1, Nygren PJ1, Purkey AM3, Colledge M4, Langeberg LK1, Dell’Acqua ML3, Zweifel LS2, Scott JD5.1HHMI/ University of Washington, United States;2University of Washington, United States;3University of Colorado Denver, United States;4Yellow-Scope, LLC, United States.5HHMI/ University of Washington, United States;

Abstract

Anchoring proteins direct protein kinases and phosphoprotein phosphatases toward selected substrates to control the efficacy, context, and duration of neuronal phosphorylation events. The A-Kinase Anchoring Protein AKAP79/150 interacts with protein kinase A (PKA), protein kinase C (PKC), and protein phosphatase 2B (PP2B, calcineurin) to modulate second messenger signaling events. In a mass spectrometry based screen for additional AKAP79/150 binding partners, we have identified the Roundabout axonal guidance receptor Robo2 and its ligands Slit2 and Slit3. Biochemical and cellular approaches confirm that a linear sequence located in the cytoplasmic tail of Robo2 (residues 991-1070) interfaces directly with sites on the anchoring protein. Parallel studies show that AKAP79/150 interacts with the Robo3 receptor in a similar manner. Immunofluorescent staining detects overlapping expression patterns for murine AKAP150, Robo2, and Robo3 in a variety of brain regions including hippocampal region CA1 and the islands of calleja. In vitro kinase assays, peptide spot array mapping, and proximity ligation assay staining approaches establish that human AKAP79-anchored PKC selectively phosphorylates the Robo3.1 receptor subtype on serine 1330. These findings imply that anchored PKC locally modulates the phosphorylation status of Robo3.1 in brain regions governing learning and memory, and reward.


Posted in Edition 53 - 2015, June, Subject Review
Previous Editions
Images
Instructivo para mandar imágenes
CLICK AQUI
RSS Feed