bluuebunny/crossref_metadata_2025 huggingface dataset:
FRI168 LH-hCG Receptor Is Expressed In All Three Zones Of Normal Adrenal Glands But Not In Aldosterone-Producing Micronodules
Authors: Reda Sarah Alicia , Kirschenbaum Alexander , Yao Shen , Milgram Lucas , Rege Juilee , Levine Alice C
Published: October 2023

Disclosure: S.A. Reda: None. A. Kirschenbaum: None. S. Yao: None. L. Milgram: None. J. Rege: None. A.C. Levine: None.

Embryologically, the adrenals and gonads develop from the urogenital ridge with differential migration and differentiation. Post-natally, pituitary adrenocorticotropic hormone (ACTH) stimulates adrenal cortical cell growth and steroidogenesis, whereas pituitary luteinizing hormone (LH) regulates gonadal cell growth and steroidogenesis. However, there are multiple reports of gonadal tumorigenesis driven by high ACTH (in congenital adrenal hyperplasia) and adrenal cortical tumorigenesis driven by high LH/human chorionic gonadotropin (hCG) levels (adrenal tumors in pregnant and postmenopausal females). We studied the immunohistochemical expression of LH/hCG receptor (LH/hCG-R) in normal adrenal glands from 12 kidney donors (four male and eight female, two of which were <50 years of age and six were >50 years of age). Some of these specimens also contained aldosterone producing micronodules (APMs). Staining was performed for Synaptophysin, aldosterone synthase (CYP11B2), LH/hCG-R, WNT/β-Catenin, and DLK-1. In normal adrenal tissue, we first used synaptophysin stain to identify the nerves in the adrenal cortex as well as the adrenal medulla (M). CYP11B2 staining demonstrated immunoreactivity in a linear pattern throughout the Zona Glomerulosa (ZG) in five of the 12 samples. Faint expression of LH/hCG-R was detected in all twelve samples throughout the three layers of the adrenal cortex: the ZG, Zona Fasciculata (ZF) and Zona Reticularis (ZR). However, there were occasional cells in both the ZR and ZF that stained intensely for LH/hCG-R expression. LH/hCG-R immunoreactivity was also noted in the progenitor-rich subcapsular (SC) region. WNT β-Catenin, a transcription factor involved in the proliferation of adrenal cortical cells, stained most heavily in the SC/ZG region, possibly representing adrenal cortical progenitor cells. DLK-1, an adrenal cortical progenitor cell marker, was also expressed in the subcapsular zone. In adrenal tissues containing APMs, CYP11B2 immunostaining revealed areas of CYP11B2 positive cells that dip into the ZF, with no preserved linear staining for CYP11B2 in the ZG. LH/hCG-R stained most densely in the SC and ZG regions, with little to no immunoreactivity in the APMs. WNT β-Catenin likewise stained the SC and ZG regions with no expression in the APMs. Our findings suggests that LH/hCG-R is expressed sporadically in normal adrenal cortical tissue, but densely at times in all three zones of the adrenal cortex and the subcapsular area where the putative adult progenitor cells reside. The LH/hCG-R does not appear to be expressed throughout the entirety of APMs.

Presentation: Friday, June 16, 2023
SAT357 Rising Incidence, Health Resource Utilization and Costs of Polycystic Ovary Syndrome in the United Kingdom
Authors: Berni Tom , Morgan Chris , Rees Aled
Published: October 2023

Disclosure: T. Berni: None. C. Morgan: None. A. Rees: None.

Background: Polycystic Ovary Syndrome (PCOS) is a common disorder but trends in incidence and impact on health resource utilization are not known. Objectives: 1. To describe secular trends in prevalence and incidence of PCOS in the UK. 2. To establish healthcare resource use and associated primary and secondary care costs. Methods: Data were extracted from the Clinical Practice Research Datalink Aurum database and linked Hospital Episode Statistics (HES) database (2004-2021). Patients (≥18 yrs) of acceptable research quality and eligible for linkage to HES were included. A diagnosis of PCOS was established from medcode or ICD-10 codes. Point prevalence was assessed at the midpoint of each year from 2004-2020. Incident cases (2004-2021) were defined as a first diagnosis (index date) at least 90 days after practice registration. Costs from index date were compared for incident cases (n=132,925) and controls matched (1:1) by age, BMI category and primary care practice. Primary care contacts were assigned an average cost as listed in the Unit Costs of Health and Social Care 2019. Prescription items were assigned a net ingredient cost from the Prescription Cost Analysis for England 2019. Inpatient admissions and outpatient consultations were processed into Healthcare Resource Groups and costed to the National Tariff. Results: The incidence of diagnosed PCOS increased from 1.22 per 1000 person years in 2004 to 1.77 in 2012 and 2.20 in 2019. Point prevalence increased from 0.9% in 2004 to 2.1% in 2012 and 3.4% in 2020. Mean contacts per person year (ppy) for patients with PCOS versus controls were 0.43 vs 0.25 for inpatients (p<0.001), 3.29 vs 1.83 for outpatients (p<0.001) and 6.46 vs 4.71 (p<0.001) for primary care. Mean healthcare costs (ppy) were £748 vs £438 (p<0.001) for inpatients, £383 vs £216 (p<0.001) for outpatients, £156 vs £111 for primary care and £114 vs £87 (p<0.001) for primary care prescriptions. Total health care contacts ppy were 10.81 vs 6.21 (p<0.001) and total associated costs were £1400 vs £852 (p<0.001). Conclusion: The prevalence and incidence of diagnosed PCOS has risen significantly in the United Kingdom over the last two decades. This may be partly due to increased ascertainment and recording on computerised systems. Health resource utilization and costs were almost doubled in women with PCOS compared to controls.

Presentation Date: Saturday, June 17, 2023
SAT367 A Rare Case of Ovarian Cellular Fibroma Producing Testosterone
Authors: Rai Harpreet Kaur , Dandona Paresh
Published: October 2023

Disclosure: H.K. Rai: None. P. Dandona: None.

Background: Ovarian fibromas are hormonally inactive, benign sex cord-stromal tumors. However, the occurrence of ovarian fibromas in a woman presenting with post-menopausal bleeding, hirsutism, and elevated testosterone levels raises suspicion of such tumors being functionally active.Case: 62-year-old menopausal woman presented for evaluation of worsening facial hair for the past year. She reported vaginal spotting for the past two months and was following up with a gynecologist. She denied acne, male pattern baldness, or any other virilizing features. She was also recently diagnosed with polycythemia; hematological workup was remarkable for elevated testosterone. Her blood work was significant for elevated testosterone levels and hemoglobin, results as follows: total testosterone: 160ng/dl (2-45 ng/dL), free testosterone: 25.4pg/ml (0.1-6.4 pg/ml), sex hormone binding globulin (SHBG): 26 nmol/L (14-73 nmol/L), estradiol: <15 pg/ml (post-menopausal range: </=31 pg/ml), FSH: 17.3mIU/ml (23.0-116.3mIU/ml), LH: 19.1mIU/L (10.0-54.7 mIU/mL), prolactin: 5.7 ng/mL (2.0-20.0 ng/mL), DHEAS: 78 mcg/dL (12-133 mcg/dL), 8 AM cortisol: 17 mcg/dL (10-20 mcg/dL), ACTH: 11pg/ml (6-50pg/ml), Hb: 16.6g/dl (11.7-15.5 g/dL) and hematocrit: 49.8% (35.0-45.0%). Ultrasound pelvis showed heterogeneous thickened endometrium, small left-sided intramural leiomyoma of the uterus and ovaries were not well identified. MRI pelvis with and without contrast showed a 2.4 cm left uterine fibroid. A simple cyst was noted in the right ovary and two para-ovarian cysts were noted on the left ovary. The overall appearance of both ovaries was similar to a CT scan done a year earlier. She underwent a total hysterectomy with bilateral salpingo-oophorectomy. Surgical pathology was remarkable for cellular fibroma of both ovaries. After surgery, the patient's facial hair significantly improved, and she was noted to have normal testosterone levels (total testosterone: 23, free testosterone: 2.6, SHBG: 50). Her hemoglobin normalized as well, reaching a value of 13 g/dL. Conclusion: Our patient presented with post-menopausal bleeding, hirsutism, polycythemia, and increased testosterone levels. After she underwent total hysterectomy with bilateral salpingo-oopherectomy, her symptoms resolved. Testosterone and hemoglobulin levels returned to normal. Surprisingly, ovarian cellular fibroma on both sides was identified by surgical pathology. Ovarian cellular fibromas are rarely linked to endocrine activity, but with this case, we want to emphasize that its hormonal activity needs further exploration in the future.

Presentation Date: Saturday, June 17, 2023
SAT372 Obstructive Sleep Apnea (OSA) Risk Among Patients With Polycystic Ovary Syndrome (PCOS)
Authors: Christ Jacob P , Shinkai Kanade , Corley Jamie , Pasch Lauri , Cedars Marcelle Ivonne , Huddleston Heather
Published: October 2023

Disclosure: J.P. Christ: None. K. Shinkai: None. J. Corley: None. L. Pasch: None. M.I. Cedars: None. H. Huddleston: None.

Background: Risk of OSA is increased up to seven-fold among patients with PCOS independent of body mass index (BMI). The cause of this increased risk is poorly understood. Likely a bi-directional relationship exists whereby OSA may worsen insulin resistance and hyperandrogenism which in turn may alter airway physiology. Improved understanding of the link between OSA risk and reproductive and metabolic features of PCOS could reveal new strategies to improve a variety of health outcomes for this population. Objective: To assess associations between OSA risk and reproductive and metabolic features among patients with PCOS. Methods: A cross-sectional analysis of patients that underwent a standardized evaluation for PCOS at a single tertiary center from 2017 to 2022 was completed. Patients were eligible for inclusion if they were aged 18-44, met modified Rotterdam criteria for PCOS and had completed a Berlin Questionnaire (BQ) for OSA risk assessment. The BQ contains 10 questions in 3 categories and a high-risk response was recorded if 2 out of 3 categories were positive. All patients underwent standardized anthropometric, ultrasound, endocrine, and metabolic phenotyping. PCOS features were compared between those with and without a high risk BQ. Results: Of the 572 patients screened, 309 patients with PCOS met inclusion criteria. Of these, 104 (33.7%) patients had a high risk BQ. Compared to those without a high risk BQ, those with high-risk BQ had significantly higher waist to hip ratio (WHR) (mean ± standard deviation, 0.9±0.1 vs 0.8±0.1), LDL (113±31.1 vs 103.3±26.5 mg/dL), triglycerides (132±75 vs 89±55 mg/dL), fasting insulin (22±21.2 vs 13.9±16.9 mIU/mL), 2-hour insulin (125±141.2 vs 68.4±108.9 mIU/mL), fasting glucose (100±39 vs 88±20 mg/dL), 2-hour glucose (125.7±56.4 vs 101.7±40 mg/dL), hemoglobin A1c (5.7±1.1 vs 5.2±0.6%), C-reactive protein (6.2±5.7 vs 2.8±3.8 mg/L), ALT (37±50 vs 27±27 U/L), free testosterone (6.5±3.9 vs 5.1±3.5 pg/mL), and lower HDL (49.7±12.4 vs 61.1±16.5 mg/dL) and sex hormone binding globulin (SHBG) (32.3±20.2 vs 53.7±37 nmol/L); p<0.05 for all comparisons. In multivariable regression, controlling for WHR and age, C-reactive protein (β=0.095, p=0.005), SHBG (β=-0.014, p=0.037), HDL (β=-0.025, p=0.032) and Hemoglobin A1c (β=0.421, p=0.073) were found to independently predict presence of high risk BQ. Conclusions: High risk for OSA is a frequent finding among patients with PCOS. Risk for sleep apnea appears to be associated with markers of dysglycemia, dyslipidemia, inflammation, and hyperandrogenism independent of adiposity. Future research is needed to determine if treatment of OSA has metabolic and/or reproductive benefits.

Presentation Date: Saturday, June 17, 2023
FRI173 Investigation Of Cut Off Values For Identifying Cortisol Excess In Adrenal Venous Blood Sampling
Authors: Watanabe Kazuki , Nakai Kazuki , Tsuru Tani Yuya , Hirose Rei , Saito Jun , Nishikawa Tetsuo
Published: October 2023

Disclosure: K. Watanabe: None. K. Nakai: None. Y. Tsurutani: None. R. Hirose: None. J. Saito: None. T. Nishikawa: None.

Objective: Primary aldosteronism (PA) is sometimes accompanied by subclinical Cushing’s syndrome (SCS). However, there is no defined cutoff value for excess cortisol (Cort) in adrenal venous blood sampling (AVS). Meanwhile, some reports have shown AVS usefulness in overt Cushing’s syndrome. This study analyzed AVS parameters to determine the cutoff value of excess Cort in AVS. Methods: The Cort-producing adenoma (CPA) group included patients with adrenal tumors who had a Cort level of ≥ 5 mg/dL after a 1 mg dexamethasone suppression test (DST) and underwent segmental AVS (sAVS) at our hospital from 2009-2020 (n = 75). Patients who underwent AVS on only one side had an adrenocorticotropic hormone (ACTH) level of > 10 pg/mL after 1 mg DST, a midnight Cort level of < 5.0 mg/dL, bilateral cortisol-producing tumors, and/or adrenal cancer, were excluded (n = 26). The control group with normal Cort secretion included patients with Cort level of < 1.8 mg/dL after 1 mg DST who underwent sAVS and whose resected adrenal tumor was negative or weakly positive in 11β-hydroxylase cytochrome P450 staining (n = 45). We compared the various sAVS values between the CPA and control groups. Results: Forty-nine patients in the CPA and 45 in the control groups were analyzed. Compared with the control, the CPA group had a significantly higher maximum Cort level in the tributary veins flowing from the tumor (1220 [860.1-1584.4] vs. 928 [695.5-1350.0], p = 0.016). Meanwhile, Cort levels in the central vein on the tumor side were not significantly different between the two groups (895 [614.7-1312.7] vs. 820 [590.5-1065.0], p = 0.202). The CPA group had a significantly lower minimum Cort level in all tributary veins (117 [73.1-241.5] vs 559 [386.0-794.0], p < 0.001), and Cort level in the central vein on the non-tumor side (176 [93.9-394.0] vs 766 [499.5-1090.0], p < 0.001). The ratio of each value showed that the maximum Cort level in tributary veins flowing from the tumor/minimum Cort level in all tributary veins (9.59 [3.75-21.23] vs 1.65 [1.45-2.06], p < 0.001) and Cort level in the central vein on the tumor side/Cort level in the central vein on the non-tumor side (4.44 [1.95-11.35] vs 1.04 [0.73-1.34], p < 0.001) were significantly higher in the CPA group. The former had the largest area under the curve for the receiver operating characteristic curve in various indices (AUC = 0.948, 81.6% sensitivity and 97.8% specificity when 3.28 was set as the cut-off value). Conclusion: Novel cutoff values were proposed to identify excess cortisol in AVS.

Presentation: Friday, June 16, 2023
SAT379 Potassium Levels in Women with PCOS Using Spironolactone for Long-term
Authors: Oliveira Thais Areias , Marchesan Lucas Bandeira , Spritzer Poli Mara
Published: October 2023

Disclosure: T.A. Oliveira: None. L.B. Marchesan: None. P.M. Spritzer: None.

Introduction. Polycystic Ovary Syndrome (PCOS) is the most common endocrine disorder among women of reproductive age. Spironolactone (SPL) has been used off-label to manage hyperandrogenic manifestations in women with PCOS but the incidence of hyperkalemia in these population is scarce. Materials and Methods. In this retrospective study, we assessed data collected from electronic medical records of women with PCOS that received spironolactone, in addition to other drugs, to manage hyperandrogenic manifestations. Women with PCOS, defined by the Rotterdam criteria, without heart or kidney disease, who were treated with SPL for a minimum of 12 months and had records of serial potassium concentrations, were included for the analyses. Mild hyperkalemia was defined as potassium concentration between 5.1 to 5.5mEq/L, moderate 5.6 to 6,0mEq/L or severe ≥ 6,0mEq/L. Results. Data from 78 women with PCOS with potassium dosages totaling 98 measurements, were analyzed. Mean age was 29.1 ± 9.6 years, BMI 32.2 ± 8.1 kg/m², 9 had Diabetes (9.2%) and 22 (22.4%) pre-diabetes. SPL was used in association with combined oral contraceptives (COCP) in 55 participants (56.1%), progestin only pill (POP) in 21 (21.4%), metformin (MTF) in 35 (35.7%) and ACEi/angiotensin receptor blockers (ARBs) in 15 (15.3%). The median SPL dose was 100mg (50 to 150mg). Ten(12,8%) women had hyperkalemia, 4 of them more than once during follow-up, all classified as mild. The most common minor adverse effects included menstrual irregular bleeding in 8 (8.2%) in 5 using POP irregularly and 3 using COCP, followed by dizziness in 6 (6.1%) and postural hypotension in 3 (3.1%). Only in two cases (2.1%) SPL was discontinued due to poor tolerance, one due to breast pain and the other due to cramps. Conclusion: The present study suggests that women with PCOS, without kidney or heart diseasee, using SPL associated with hormone contraception for managing clinical hyperandrogenism, have a low incidence of hyperkalemia, no life-threatening episodes, well-tolerated minor adverse effects and a low rate of drug discontinuation. Sources of research support: INCT/CNPq and FAPERGS.

Presentation Date: Saturday, June 17, 2023
SAT434 Estradiol Controls GnRH-II Gene Expression In The Hypothalamus And Pituitary Gland Of Female Rhesus Macaques (Macaca Mulatta)
Authors: Lomniczi Alejandro , Luisa Appleman Maria , Urbanski Henryk F
Published: October 2023

Disclosure: A. Lomniczi: None. M. Appleman: None. H.F. Urbanski: None.

It has been known for many years that rhesus macaques express two forms of gonadotropin-releasing hormone (GnRH-I and GnRH-II) in the medial basal hypothalamus (MBH), and that both forms are capable of stimulating the release of luteinizing hormone (LH) in vivo. However, while much has been published about the role of GnRH-I in reproduction, very little is known about the function of GnRH-II. To shed light on this issue, we studied the expression pattern of these two genes in different parts of the monkey hypothalamus, median eminence and pituitary gland. Quantitative PCR (qPCR) analysis revealed that while GnRH-I is preferentially expressed in the Arcuate Nucleus (ARC), Median Eminence (ME), Supraoptic Nucleus (SON) and Suprachiasmatic Nucleus (SCN); GnRH-II was highly expressed in the SON, ARC-ME and Paraventricular Nucleus (PVN), confirming our previously published in situ hybridization studies. Strikingly, GnRH-II mRNA was also detected in the pituitary gland, a site where GnRH-I was not found. We have previously demonstrated that maximum hypothalamic GnRH-II expression occurs around the time of the preovulatory LH surge, when estrogen (E2) levels are maximal, but no significant changes in GnRH-I mRNA levels occur. Here using an ovariectomized (OVX) macaque model subjected to E2 replacement paradigm, designed to mimic the plasma E2 levels observed during the preovulatory estradiol peak, we performed RNAseq and qPCR from the PVN and pituitary gland to identify genes differentially regulated by E2. While GnRH-I mRNA was not affected by E2 treatment in any of the tissues studied, pituitary GnRH-II expression highly correlated with estrogen levels (r>0.8; p<0.01), with over a hundred-fold increase in mRNA expression. Taken together, these data suggest that pituitary GnRH-II (but not GnRH-I) gene expression is induced by a positive feedback action of estradiol. This novel finding raises the possibility that GnRH-II plays a major role in triggering the preovulatory LH surge in primates, not only at the ARC-ME but also at the pituitary level.

Presentation Date: Saturday, June 17, 2023
OR35-03 Uncovering Causative Mutations in Polycystic Ovary Syndrome
Authors: Srivastava Jaya , Ovcharenko Ivan
Published: October 2023

Disclosure: J. Srivastava: None. I. Ovcharenko: None.

PCOS is a complex disorder manifesting as reproductive and metabolic abnormalities in women. The undefined etiology stems from our incomplete understanding of genetic aberrations contributing to the diseased phenotype. This has resulted in speculations surrounding the cause-consequence relationship between phenotypic features of PCOS and limited our knowledge of comprehensive genetic signatures of PCOS. Pleiotropic effects are salient features of genetic regulation in mammalian genomes. The contribution of a set of regulatory elements (REs) to upregulation of a single gene, or the genetic control of multigene loci by a single RE, give rise to heterogenous phenotypes. Enhancers in particular dictate the spatio-temporal scale of gene expression by controlling transcription factor (TF) binding events. The complex traits of PCOS are suggestive of pleiotropy originating from genetic and epigenetic changes that arise from nucleotide variations in REs of the genome. Genome Wide Association Studies (GWAS) have identified ∼100 risk variants associated with PCOS and implicated the involvement of several genes such as FSHB/R, THADA, DENND1A, etc. However, mechanisms by which they contribute to pathophysiology are unknown. Additionally, twin studies have implicated the heritability of PCOS to be ∼80% and yet, the heritability from GWAS variants accounts for less than 10%. Experimental data may account for the ‘missing heritability problem’ specifically through rare variants that are missed in GWAS. To address this issue, we developed a Deep Learning model of gene regulation in PCOS identifying hundreds of active REs orchestrating the activity of PCOS genes. We also established a foundation for quantifying the impact of single-nucleotide noncoding mutations on TF binding and RE activity. By combining thousands of experimental assays associated with a large number of biological events such as tissue-specific epigenetic maps of the human genome, multi-tissue profiling of TF binding and gene expression using data from high throughput experiments we devised a strategy to accurately identify functionally map causative mutations in the GWAS loci of PCOS genes. MPRA experimental data validates a large fraction of our predicted causative variants thus suggesting that our studies have a potential of a leap forward accurate dissection of genomic and epigenetic mechanisms of PCOS. Consolidated results of this analysis and experimental validation of our predictions will lead to detailing the regulatory mechanisms of PCOS.

Presentation Date: Sunday, June 18, 2023
FRI162 Ligand-dependent Photocrosslinking of Mineralocorticoid Receptors Using Site-directed Incorporation of Unnatural Amino Acids
Authors: Almeida-Prieto Brian , Gonzalez-Hernandez Alberto J , Giraldez Teresa , Alvarez de la Rosa Diego
Published: October 2023

Disclosure: B. Almeida-Prieto: None. A.J. Gonzalez-Hernandez: None. T. Giraldez: None. D. Alvarez de la Rosa: None.

Steroid receptors dimerize prior DNA binding as a key step for translational activation of target genes. It appears that several domains of the receptors play a role in the process. In addition, current evidence suggests that the active conformation of steroid receptors may not always be a dimer, but involves different quaternary structures, characteristic for each receptor. Recent data obtained by our group suggests that agonist-bound mineralocorticoid receptors (MR) adopt a tetrameric form in the nucleoplasm, forming higher order oligomers when bound to hormone response elements in the chromatin. The formation of such complexes would likely requires more than one oligomerization interface. To generate a tool for investigating inter-subunit contacts, we incorporated photoactivatable crosslinkers into specific sites in MR using a genetically-encoded unnatural amino acid (p-Benzoyl-L-phenylalanine, BzF). We created a library of 26 amber stop codon mutants substituting conserved A, F, W and Y residues distributed along the N-terminal domain (NTD), DNA-binding domain (DBD) and ligand-binding domain (LBD) of the mouse MR coding sequence. Site-directed incorporation of BzF into MR was performed by co-expressing an orthogonal pair of BzF-specific aminoacyl-tRNA synthetase and an amber-suppressor tRNA with the mutant receptors. Characterization of the library showed that 18 mutants produced functional receptors after BzF incorporation. Using an in silico approach we identified position W914 in helix 10 of the LBD as a potential candidate for photocrosslinking. Treatment of cells expressing W914BzF with UV light selectively increased aldosterone-bound, but not cortisol-bound, MR transcriptional activity by approximately 50%. Increased activity was still apparent after hormone washout, suggesting that BzF-induced photocrosslinking at position 914 contributes to stabilizing an active conformation and identifying H10 as a putative region for ligand-specific oligomerization interface. Our results support the strong potential of this methodology for studying this new organization model proposed for MR and other steroid receptors, along with the characterization of functional protein-protein interactions.

Presentation Date: Friday, June 16, 2023
OR11-02 Structural Determinants of Pure Antiestrogenic Activity
Authors: Vallet Amandine , Diennet Marine , Thiombane Khady , Vivet Anaïs , Weber Sandra , El Ezzy Mohamed , Shaikh Faraz , Poupart Julien , Mendoza-Sanchez Rodrigo , Alexandra Schuetz Doris , Marinier Anne , Greene Geoffrey L , William Fanning Sean , Nathalie Mader Sylvie
Published: October 2023

Disclosure: A. Vallet: None. M. Diennet: None. K. Thiombane: None. A. Vivet: None. S. Weber: None. M. El Ezzy: None. F. Shaikh: None. J. Poupart: None. R. Mendoza-Sanchez: None. D. Schuetz: None. A. Marinier: None. G.L. Greene: None. S.W. Fanning: None. S.N. Mader: None.

Inhibitory activities of antiestrogens on estrogen receptor alpha (ERα) range from mixed antagonism/agonism (selective ER modulators; SERMs) to complete antiestrogenicity associated with accelerated ERα turnover (selective ER degraders; SERDs). Using a panel of SERMs, SERDs and PROTACs, we show that antiestrogens induce variable degrees of SUMOylation of ERα and that efficient induction of SUMOylation, rather than increased suppression of coactivator recruitment or accelerated ERα degradation, correlates with suppression of ERα basal and induced transcriptional activity in reporter assays. Antiestrogens did not induce SUMOylation of ERβ and did not suppress its basal transcriptional activity in the same assays. Chimeras between the two receptors identified ERα-specific residues required for induction of SUMOylation by antiestrogens and revealed differences according to the antiestrogen structure. In addition, induction of ERα SUMOylation by antiestrogens depended on the hydrophobicity of N-terminal residues of ligand binding domain (LBD) helix H12. L536 mutations, including those occurring in endocrine therapy-resistant breast cancer, abolished both induction of ERα SUMOylation and pure antiestrogenicity. Structures of the L536S human ERα LBD bound to fulvestrant analogs and molecular dynamics simulations revealed dynamic side chain interactions with ERα H12 in the coactivator-binding groove, and predict the impact of SUMOylation-suppressing L536 mutations on these interactions. Together, our results show that antiestrogen-induced ERα SUMOylation is associated with complete suppression of its transcriptional activity and provide insights into the structural determinants of pure antiestrogenicity.

Presentation Date: Friday, June 16, 2023
SAT020 Is Testosterone The Male Sex Hormone Responsible For Increased Male Mortality In Melanoma? In-vitro Studies On Two Human Melanoma (BLM, 1205Lu) Cell Models
Authors: Ramaraj Pandurangan
Published: October 2023

Disclosure: P. Ramaraj: None.

Epidemiological data pointed to increased male mortality in melanoma and clinical studies pointed to better protection of menstruating females in melanoma than post-menopausal women and men of any age. Our in-vitro studies with progesterone (P) on human melanoma (BLM and 1205Lu) cell lines showed a significant decrease in cell growth along with a decrease in the secretion of proinflammatory cytokine IL-8 by these cells. These in-vitro studies correlated with the hormonal data which showed that progesterone levels in menstruating females ranged between 1000 - 1500 ng/dl, compared to post-menopausal women 20 - 100 ng/dl and men 27-90 ng/dl, suggesting low progesterone levels meant no protection in melanoma. This observation prompted us to hypothesize that lack of progesterone in males could be a cause for increased male mortality. This hypothesis also raised the question of the role of male sex hormones (AD-androstenedione and T-testosterone) in increased male mortality. In order to address these questions in-vitro experiments were planned. First, to indirectly induce endogenous IL-8 by pretreating the cells with endothelin and then adding steroid hormones (AD, T, P). Secondly, to add IL-8 directly to the cells and then add steroid hormones. Finally, to suppress endogenous IL-8 with curcumin and then add T to one set of cells and IL-8 + T to another set of cells. For the first experiment, the result was a marginal increase in cell growth (67%) in cells pretreated with endothelin followed by T-treatment compared to straight T-treated cells (60% cell growth) without endothelin pretreatment. For the second experiment, there was a slight increase in cell growth in IL-8-added cells, but it was not statistically significant. For the final experiment, the cells treated with IL-8 + T after curcumin pretreatment showed a partial restoration of cell growth (49%) compared to cells (40% cell growth) treated with plain T after curcumin treatment, suggesting T was not able to control or regulate IL-8 action like progesterone. These in-vitro experiments suggested that in males 2 reasons 1) a deficiency of progesterone and 2) inefficiency of T to control endogenous IL-8 could possibly result in increased IL-8 levels. This increased IL-8 level in males could lead to increased cell growth and metastasis in melanoma leading to death. So, a situation arising out of male reproductive endocrine physiology could probably be responsible for increased male mortality in melanoma.

Presentation: Saturday, June 17, 2023
OR22-05 Progesterone Receptor-Grb2 Interaction Is Associated With Better Outcomes In Breast Cancer
Authors: Wittayavimol Nattamolphan , Iwabuchi Erina , Pateetin Prangwan , Miki Yasuhiro , Onodera Yoshiaki , Sasano Hironobu , Boonyaratanakornkit Viroj
Published: October 2023

Disclosure: N. Wittayavimol: None. E. Iwabuchi: None. P. Pateetin: None. Y. Miki: None. Y. Onodera: None. H. Sasano: None. V. Boonyaratanakornkit: None.

The presence of progesterone receptor (PR) is usually associated with better clinical outcomes for several endocrine-related cancers. However, the molecular mechanism of how PR mediates growth inhibition remains unclear. In addition to its role as a nuclear transcription, PR also possesses an extranuclear function, mediated mainly through PR polyproline domain (PPD) interaction with the SH3 domains of cytoplasmic signaling molecules. Our previous study suggested a possible role of PR-PPD in mediating SH3-domain interaction. We showed that PR-PPD and SH3 domain interaction inhibited EGF-mediated signaling and decreased lung cancer cell proliferation. An essential adapter protein that helps transduce growth factor signaling is Grb2 which has an SH2 domain flanked by two SH3 domains. In this study, we hypothesized that PR interferes with cytoplasmic signaling through interaction between PR-PPD and Grb2-SH3. We used several protein-protein interaction assays in vitro and in cells to demonstrate PR-Grb2 interaction. GST-pulldown analysis showed that PR-PPD interacts specifically with SH3 domains of Grb2. No interaction was detected when key prolines in the PR-PPD to alanines. Immunofluorescence staining showed colocalization of PR and Grb2 in both the nucleus and cytoplasm of BT474 breast cancer cells. Using Bimolecular Fluorescence Complementation (BiFC) analysis, we showed that PR and Grb2 interacted in breast cancer cells, specifically through the Grb2-SH3 domain. No BiFC signal was detected when Grb2-SH3 domains were deleted. Furthermore, we used Proximity Ligation Assay (PLA) analysis to examine protein-protein interaction of a distance less than 40nm in cells. We found PR-Grb2 interaction in PR-positive breast cancer tissue sections. No PLA signal was detected in PR-negative breast cancer cells. Importantly, we examined 43 PR-positive breast cancer specimens and showed that PR-Grb2 interaction was associated with low histological stage and negatively correlated with lymph node invasion and metastasis of breast cancer. Our results provide a molecular mechanism for how PR can inhibit growth factor signaling by interfering with Grb2 signaling and a basis for developing novel therapeutic strategies for future cancer treatment.

Presentation: Saturday, June 17, 2023
OR33-01 Live Cell Imaging Of Enhancer Transcriptional States
Authors: Chapus Fleur , Day Christopher , Yasar Pelin , Bregio Celyn , Palia Alyson , Scappini Erica , Tucker Charles , Rodriguez Joseph
Published: October 2023

Disclosure: F. Chapus: None. C. Day: None. P. Yasar: None. C. Bregio: None. A. Palia: None. E. Scappini: None. C. Tucker: None. J. Rodriguez: None.

Intra-tumoral gene expression heterogeneity of estrogen receptor positive breast cancers is a roadblock toward effective treatment due to subpopulations acquiring resistance and metastatic ability. This expression heterogeneity can be explained by the stochastic nature of transcription. Transcriptional initiation is coordinated through distal elements known as enhancers. These elements aid in the recruitment of cofactors and chromatin remodelers to promoters. During transcriptional activation, genes are transcribed during short periods of nascent RNA synthesis known as transcriptional bursts. Furthermore, estrogen responsive genes can transition between transcriptional bursting and variably long periods of inactivity lasting from minutes to days. Here, we investigated whether estrogen responsive enhancers of variably expressed genes also exhibited transcriptional states which reflect promoter states. To observe live cell transcription dynamics, we integrated MS2 stem loops into several endogenous enhancers using CRISPR. After transcription initiation, these stem loops are transcribed and subsequently bound by the RNA binding protein, MS2-GFP. This enrichment in MS2-GFP at the transcription site is visible as a bright punctate spot. We observed that the estrogen responsive TFF1 enhancer exhibited short periods of transcriptional activity and long periods of inactivity. Moreover, the TFF1 enhancer RNA (eRNA) sporadically formed dynamic, but rare and long-lived condensates. To correlate enhancer and promoter transcription we used single molecule fluorescence in situ hybridization of several eRNA. We observed that eRNA aggregates were correlated with an increase of mRNA content and occasionally localized to different nuclear bodies. Lastly, we enriched for chromatin bound RNA and observed long enhancer RNA transcripts. Taken together, our work highlights the dynamic coupling of enhancer and promoter regulation and the importance of understanding the regulators of enhancer transcriptional states in cancer.

Presentation: Sunday, June 18, 2023
THU653 Subacute Thyroiditis Following Covid19 Infection
Authors: Fanous Ereeny , Georges Karam Jocelyne
Published: October 2023

Disclosure: E. Fanous: None. J.G. Karam: None.

Background: Subacute thyroiditis is a self-limiting inflammatory condition of the thyroid gland which usually occurs as a post-viral reaction, leading to the destruction of thyroid follicular cells due to infiltration of lymphocytes. During the first outbreak of COVID-19 virus, subacute thyroiditis had been reported as a complication that may occur in patients without any previous thyroid disorder [1]. Clinical Case: A 62-year-old female patient presented with few weeks history of generalized weakness, fatigue, chills, malaise, low appetite, and unintentional weight loss of around 10lbs. She stated that she feels "internal tremors" and palpitations with heart rate increasing from baseline 60 bpm to 90bpm. She had one episode of near syncope at work prompting her presentation. She reports no change in bowel movements, no heat intolerance, no fever or night sweat, no shortness of breath, no visual changes or eyes problems. Five weeks prior to presentation, the patient was diagnosed with COVID-19 infection by PCR, with a mild disease pattern that consisted of upper respiratory symptoms lasting for few days, however she continued to feel fatigue and low appetite since then, attributing her symptoms to long-term sequelae of Coivd-19. Her past medical history is remarkable for obesity, hypercholesterolemia, and two prior Covid 19 infections: in 2020, treated with monoclonal antibodies, and early 2022. The physical exam was significant for slightly tender thyroid and heart rate of 94 bpm. Thyroid function tests were consistent with hyperthyroidism; TSH<0.01mIU/ml, FT4=2.55ng/dl (0.58-1.64), T4=19.6 mcg/dl (4.4-9.5), T3=1.45ng/ml (0.72-1.35), with negative TSI, TPO AB, and TG AB. ESR was 44mm/hr, RAI uptake and scan showed very low uptake which is indicative for subacute thyroiditis.

The patient was given supportive therapy. She developed mild and transient hypothyroidism at 6 weeks follow-up with TSH of 8.59 IU/ml and FT4=0.62 ng/dl (0.89-1.76) before her thyroid tests return to normal at 9 weeks with complete resolution of her symptoms. Conclusion: Prolonged diverse symptoms following Covid19 infection are often attributed to long term Covid symptoms. Health Care providers should be aware of the possibility of subacute thyroiditis following Covid 19 infection and should obtain thyroid function tests in patients who present with malaise, decreased appetite, weight loss and palpitations in the weeks following Covid 19 infection. Reference:1. Mattar SAM, Koh SJQ, Rama Chandran S, Cherng BPZ. Subacute thyroiditis associated with COVID-19. BMJ Case Rep. doi: 10.1136/bcr-2020-237336

Presentation: Thursday, June 15, 2023
THU656 Subacute Thyroiditis In Early Pregnancy: A Rare Case Report
Authors: El Sayed Farah , Shelden Daniel
Published: October 2023

Disclosure: F. El Sayed: None. D. Shelden: None.

Background Subacute thyroiditis (SAT) is a self-limiting inflammatory disorder that can be caused by a viral infection and presents with neck pain, fever, and fatigue. To date, only 8 cases of SAT have been reported in pregnant women. Hyperthyroidism during pregnancy is associated with both maternal and neonatal complications. The early diagnosis and treatment of SAT during pregnancy may play a vital role in prevention of adverse fetal outcomes. Case Presentation A 37-year-old female in her 6th week was referred to the endocrinology clinic for neck pain that started two months ago following a viral sore throat. She also reported exhaustion, palpitations, and occasional dysphagia. This is her 10th pregnancy, and she reports a history of thyroid isthmic cysts with hemorrhagic degeneration drained in 2014.The patient is a former occasional smoker and denies alcohol or drug use. Family history is significant for autoimmune disease in her sibling, and type 2 diabetes mellitus in her father. On physical exam, her neck is soft, supple, with no lymphadenopathy. The thyroid gland is enlarged to approximately 40 grams and is mildly tender to palpation. Labs were significant for thyroid-stimulating hormone (TSH) of 0.01 uIU/ml (reference range 0.4-4.5 uIU/ml), serum-free triiodothyronine (FT3) of 4.7 pg/ml (reference rage 1.7-3.7 ng/dl), serum-free thyroxine (FT4) 1.6 pg/ml (reference range 0.7-1.5 pg/ml), ESR 67 mm/hr (reference range 0-18 mm/hr), CRP 52.6 mg/L(reference range <8 mg/L). Thyroid ultrasound revealed enlarged heterogeneous thyroid gland , with poorly defined hypoechoic areas. Based on patients’ symptoms, physical examination, and laboratory findings she was diagnosed with SAT. After review of prior literature and coordination with the patient’s obstetrician, she was started on prednisone 20 mg with a plan to taper by 5 mg weekly with close observation of thyroid function tests. Antibody assessment of thyroid was diffusely negative. Repeat assessment of the patient’s thyroid function tests showed improvement in tsh, FT3, and FT4 to 0.04, 2.4, and 0.9 respectively. The patient reported feeling significantly improved at the 2 week mark. Discussion and conclusion SAT during pregnancy is rare but requires careful management. To date, various treatment modalities are reported in case studies but there is a lack of consensus on medication dosing given the rarity of the presentation. The pathogenesis of SAT is still unknown but is thought to be secondary to destruction of thyroid follicular cells, resulting in the release of thyroid hormone and symptoms of thyrotoxicosis. SAT diagnosis is based on the patient's medical history, symptoms, physical and laboratory findings, and exclusion of other reasons for thyroid dysfunction. Pregnant women with hyperthyroidism need to be differentiated from Graves' disease, and monitoring of thyroid hormone levels is important for fetal outcomes.

Presentation: Thursday, June 15, 2023
THU661 Marine-Lenhart Syndrome: An Unusual Presentation Of Hyperthyroid Disease
Authors: Ji Beisi , Adeniran Omolara , Soni Lina
Published: October 2023

Disclosure: B. Ji: None. O. Adeniran: None. L. Soni: None.

Background: Marine-Lenhart syndrome, also known as nodular Graves’ disease, is defined as the coexistence of functioning thyroid nodules and Graves’ disease. It is a rare syndrome, with a prevalence of approximately 0.8-2.7%. The two diseases may coexist or present at different times [1]. Treatment options include high doses of antithyroid medications or radioactive iodine and thyroidectomy in selected cases [2]. Clinical Case: A 72-year-old female with a medical history of right-sided breast cancer treated with lumpectomy and radiation therapy in 2012 on anastrozole was referred for multinodular goiter (MNG) evaluation in 2018. She was clinically and biochemically euthyroid. Her thyroid ultrasound demonstrated two isoechoic right lobe nodules measuring 2.1cm and 1.3cm, respectively. An isoechoic rounded isthmus nodule measures 2.1cm. Three isoechoic nodules in the left lobe measuring 2.6cm, 2.2cm, and 1.7cm. She underwent fine needle aspiration (FNA) of two nodules in 2019, and both demonstrated benign follicular nodules with Bethesda II classification. At a follow-up visit in 2021, she reported unintentional weight loss but denied any other symptoms of hyperthyroidism, including heat intolerance, palpitation, tremor, or diarrhea. Physical examination was significant for exophthalmos, lid retraction, and diffusely enlarged thyroid with non-tenderness to palpation. Thyroid function tests revealed subclinical hyperthyroidism with TSH of <0.01uIU/mL (0.27-4.2uIU/ml) and FT4 of 1.5ng/dL(0.9-1.8ng/mL). Thyroid stimulating immunoglobulin (TSI) was 3.98IU/L (0.00-0.55IU/L), and TSH receptor antibody was 7.64 IU/L (0.00-1.75 IU/L). She was started on low-dose methimazole therapy. Thyroid uptake scan showed markedly elevated radioactive iodine uptake with 63.46% in 24 hours and focal nodular regions of increased uptake of the bilateral thyroid lobes compatible with toxic multinodular goiter. Repeated thyroid ultrasound in 2022 revealed stable-sized nodules with at least two new sub-centimeter nodules in both lobes. The patient was also seen by ophthalmology and plans to start teprotumumab treatment. At her most recent follow-up visit in January 2023, she has been continued on methimazole 2.5mg daily.

Conclusion: The criteria for diagnosing Marine-Lenhart syndrome have not been well established. The possibility of Marine-Lenhart syndrome should be considered in hyperthyroid patients with positive antibodies and nodular goiter. In this case, she was initially asymptomatic with an enlarged multinodular goiter; three years later, she developed subclinical hyperthyroidism due to Graves’ disease and toxic multinodular goiter. Her symptom and thyroid function are well-controlled with low-dose methimazole.

Presentation: Thursday, June 15, 2023
THU663 Therapeutic Plasma Exchange In Refractory Atrial Fibrillation Secondary To Thyroid Storm.
Authors: Mohammed ALShehri Sara , Salem Bafadel Marwah , Aljohani Naji J
Published: October 2023

Disclosure: S.M. ALShehri: None. M.S. Bafadel: None. N.J. Aljohani: None.

Thyrotoxic crisis (thyroid storm) is a severe form of hyperthyroidism. It has a wide range of symptoms, including cardiovascular manifestations that can be life-threatening and require prompt management. Cardiac manifestations of thyroid storm include hypertension, tachycardia, congestive heart failure, cardiac ischemia, and atrial fibrillation. Standard therapeutic approaches of thyroid storm are based on the use of thionamides, corticosteroids, and nonselective beta-blockers to inhibit thyroid hormone synthesis and release while also blocking thyroxin's peripheral effects. This approach is an effective measure in the management of these manifestations. However, when a patient fails to respond to conventional therapy or there is end organ damage and surgery is not feasible, therapeutic plasma exchange (TPE) is a life-saving measure. Here, we report a case of a 32-year-old female who is known to have hyperthyroidism, but she was not compliant with her medication. She presented with symptoms of hyperthyroidism and was found to have thyroid storm with refractory atrial fibrillation. Despite the effort to control her symptoms, she developed cardiac arrest, after 15 minutes of cardiopulmonary resuscitation she revived. Amiodarone, beta blockers, and cardioversion have all failed to improve the patient's status, and her condition has deteriorated. So, we decided to use TPE, and she received three sessions in total. After the first session of TPE, her rhythm converted to sinus rhythm, and her heart rate was controlled to less than 100 beats per minute. She showed dramatic improvement clinically and biochemically. In conclusion, cardiac complications are potentially lethal complications of thyroid storm. Prompt restoration of a normal thyroid state can reverse these complications. When conventional therapy fails to ameliorate symptoms or organ deterioration is rapid and severe, TPE can be a safe and effective measure.

Presentation: Thursday, June 15, 2023
THU665 Unilateral Graves’ Disease In A Bilobar Thyroid Gland: Unusual Cause Of Hyperthyroidism
Authors: Hamid Alaybaa Khaled , Alhuzaim Omar
Published: October 2023

Disclosure: K.H. Alaybaa: None. O. Alhuzaim: None.

Introduction: Graves' disease (GD) is an autoimmune condition that affects both thyroid gland lobes. GD patients with unilateral thyroid scan uptake are rare. These patients present with classic Graves' symptoms, occasionally with Graves' ophthalmopathy characteristics and increased thyroid hormones, and scintigraphic assessment reveals radiotracer uptake in only one lobe of the thyroid gland. Patient Findings: In this case, a 48-year-old female presented with symptoms and signs indicative of thyrotoxicosis. Lab results revealed that thyroid-stimulating hormone (TSH) was undetectable, while free thyroxine (T4) and free triiodothyronine (T3) were in the high-normal range. Positive results were observed for both thyroid-stimulating immunoglobulin and TSH receptor antibodies. Ultrasonography of the thyroid gland revealed an enlarged left lobe with heterogeneous echotexture and a hyperemic gland without any focal lesions, distinct nodules, or cysts. The NM TC-99M thyroid scintigraphy revealed enlarged and increased radioisotopes uptake in the left thyroid lobe and decreased uptake in the right thyroid lobe, with no hot or cold nodules. The calculated thyroid uptake is 4.5 percent (normal range: 0.5% to 3.5 percent). The most likely diagnosis was unilateral GD. The patient was administered carbimazole and within eight weeks had improved clinically and biochemically. Summary: This patient has thyrotoxicosis symptoms and positive autoantibodies. Tc99m uptake was increased in the left thyroid lobe and decreased in the right thyroid lobe. The left thyroid lobe was hypervascular and enlarged on ultrasonography, indicating the presence of unilateral GD. Unilateral GD is a rare disease with a poorly understood pathophysiology.

Presentation: Thursday, June 15, 2023