Cortisone Acetate Research Articles

Proliferation capability of natural killer cells upon cytokines stimulation correlated negatively with serum lactate dehydrogenase level in coronary artery disease patients.

Natural killer (NK) cells are proposed to participate in coronary artery disease (CAD) development. However, little is known about how CAD patients' NK cells respond to different stimulatory factors in terms of proliferation capability. Twenty-nine CAD patients' peripheral blood NK cells were isolated and individually treated with IL-2, IL-12, IL-15, IL-18, IL-21, cortisone acetate, hydrocortisone, or ascorbic acid for 36 hours, followed by cell cycle analysis using flow cytometry. The ratio of S and G2/M phase cell number to total cell number was defined as a proliferation index (PrI) and used for proliferative capability indication. The results showed that these eight factors resulted in different life cycle changes in the 29 NK cell samples. Remarkably, 28 out of 29 NK cell samples showed an obvious increase in PrI upon ascorbic acid treatment. The serum lactate dehydrogenase (LDH) level of the 29 CAD patients was measured. The results showed a negative correlation between serum LDH level and the CAD patients' NK cell PrI upon stimulation of interleukins, but not the non-interleukin stimulators. Consistently, a retrospective analysis of 46 CAD patients and 32 healthy donors showed that the circulating NK cell number negatively correlated with the serum LDH level in CAD patients. Unexpectedly, addition of LDH to NK cells significantly enhanced the production of IFN-γ, IL-10 and TNF-α, suggesting a strong regulatory role on NK cell's function. Ascorbic acid could promote the proliferation of the CAD patients' NK cells; LDH serum level may function as an indicator for NK cell proliferation capability and an immune-regulatory factor.

Conventional steroids vs. dual-release hydrocortisone on metabolic, cardiovascular and bone outcomes in adrenal insufficiency: a 10-year study.

Adrenal insufficiency (AI) is characterized by increased mortality compared to general population, mainly due to cardiovascular disease. Conventional glucocorticoid (GC) replacement therapy has a role in determining the increased mortality risk. Primary outcome of the current study was to evaluate the impact of 10 years of conventional GCs and DR-HC on body weight changes in treatment-naïve patients with AI. Secondary outcomes were changes from baseline to 5 and 10 years in anthropometric and metabolic profile, insulin sensitivity, cardiovascular and bone parameters. We prospectively randomised 42 patients to conventional GCs (cortisone acetate or hydrocortisone) and 44 to DR-HC (1:1). Anthropometric, metabolic, cardiovascular and bone parameters were evaluated at baseline and after 5 and 10 years of follow-up. This trial is registered at ClinicalTrials.gov NCT06260462. At 10 years of follow-up, patients with conventional GCs had significantly higher values ​​of BMI (p=0.031), waist circumference (p=0.047), systolic blood pressure (p=0.039), total and LDL cholesterol (p=0.041 and p=0.042), HbA1c (p=0.040), HOMA-IR (p=0.006), AUC2h of glucose (p<0.001), thickness of the interventricular septum in diastole and of the posterior wall (both p<0.001) and significantly lower values ​​of oral disposition index (p=0.001) and ISI-Matsuda (p<0.001), lumbar spine T score (p=0.036) and femoral neck Z score (p=0.026), compared to patients treated with DR-HC. In patients with treatment-naïve AI, 10 years of conventional GC treatment is associated with a worsening of metabolic, insulin-sensitivity, cardiac and bone outcomes, while DR-HC had no impact on them achieving a lower risk of developing comorbidities.

Asymptomatic Empty Sella Syndrome: A "New" Hypothalamic Pathology or Paraphysiological Variant.

The term empty sella refers to a shrunken or displaced (by a subarachnoid diverticulum) pituitary gland. It can be primary (genetically determined) or secondary (due to trauma/surgery/radiation). It has been reported that 50% of patients are asymptomatic, and others experience symptoms, such as headache, hypertension, or visual field defects. Few cases have an empty sella syndrome, i.e., lacking functional pituitary hormones. Diagnosis is made through NMR or CT. If asymptomatic, this condition requires no treatment; otherwise, empty sella syndrome needs hormonal replacement therapy. We examined the case of asymptomatic empty sella syndrome. A 67-year-old female patient was admitted for dilatative cardiomyopathy. She had a past medical history of arterial hypertension and right ICA endovascular repair. Blood tests demonstrated hypothyroidism, hypoadrenalism, and GH deficiency, without any signs or symptoms. NRM confirmed an empty sella, hence replacement therapy with levothyroxine and cortisone acetate was started. During a follow-up evaluation, we discovered that this biochemical profile of the patient had been known for more than a decade and never treated. Despite being exposed to stress conditions, vascular surgery and angiography, she never developed an adrenal crisis, nor has she ever been symptomatic for severe hypothyroidism. Hormonal replacement therapy was performed. The described clinical scenario is rare, as usually, empty sella syndrome presents with signs of hormone deficiency, even if asymptomatic cases have been described. Some authors suggest considering it as a hypothalamic dysfunction requiring treatment; others identify it as a paraphysiological variant. However, more cases are needed to establish a correct therapeutic strategy for these patients.

Monitoring adrenal insufficiency through salivary steroids: a pilot study.

Various glucocorticoid replacement therapies (GRTs) are available for adrenal insufficiency (AI). However, their effectiveness in restoring glucocorticoid rhythm and exposure lacks adequate biochemical markers. We described the diurnal salivary cortisol (SalF) and cortisone (SalE) rhythm among different GRTs and analysed the associations between saliva-derived parameters and life quality questionnaires. Control subjects (CSs, n = 28) and AI patients receiving hydrocortisone (HC, n = 9), cortisone acetate (CA, n = 23), and dual-release hydrocortisone once (DRHC-od, n = 10) and twice a day (DRHC-td, n = 6) collected 9 saliva samples from 07:00 to 23:00. Patients compiled Pittsburgh Sleep Quality Index, Hospital Anxiety and Depression Scale, and Addison disease-specific quality-of-life questionnaires. SalE and SalF were measured by liquid chromatography-mass spectrometry. Exposure was monitored using SalE for HC and DRHC and SalF for CA. Area under the curve (AUC) was computed. Different GRTs were compared by Z-scores calculated from saliva-derived parameters. Questionnaire results predictors were evaluated with multiple regression analysis. Compared with controls, all GRTs resulted in glucocorticoid overexposure in the morning. Hydrocortisone, CA, and DRHC-td caused overexposure also in afternoon and evening. Compared with other treatments, CA determined increased Z-score-07:00 (P < .001), DRHC-td determined increased Z-score-AUC07:00→14:00 (P = .007), and DRHC-od induced lower Z-score-AUC14:00→23:00 (P = .015). Z-scores-AUC14:00→16:00 ≥ .619 best predicted questionnaire scores. None of the GRTs mimics normal glucocorticoid rhythmicity and exposure. SalE, SalF, and Z-score may be useful markers for monitoring and comparing different GRTs. Excess glucocorticoid in early afternoon best associated with depressive symptoms and worse life and sleep quality.

Effects of Dual-Release Hydrocortisone on Bone Metabolism in Primary and Secondary Adrenal Insufficiency: A 6-Year Study.

Patients with primary (PAI) and secondary adrenal insufficiency (SAI) experience bone metabolism alterations, possibly due to excessive replacement. Dual-release hydrocortisone (DR-HC) has shown promising effects on several parameters, but bone metabolism has seldom been investigated. We evaluated the long-term effects of once-daily DR-HC on bone in PAI and SAI. Patients on immediate-release glucocorticoid therapy were evaluated before and up to 6 years (range, 4-6) after switching to equivalent doses of DR-HC, yielding data on bone turnover markers, femoral and lumbar spine bone mineral density (BMD), and trabecular bone score (TBS). Thirty-two patients (19 PAI, 18 female), median age 52 years (39.4-60.7), were included. At baseline, osteopenia was observed in 38% of patients and osteoporosis in 9%, while TBS was at least partially degraded in 41.4%. Higher body surface area-adjusted glucocorticoid doses predicted worse neck (P < .001) and total hip BMD (P < .001). Longitudinal analysis showed no significant change in BMD. TBS showed a trend toward decrease (P = .090). Bone markers were stable, albeit osteocalcin levels significantly varied. PAI and SAI subgroups behaved similarly, as did patients switching from hydrocortisone or cortisone acetate. Compared with men, women exhibited worse decline in TBS (P = .017) and a similar trend for neck BMD (P = .053). After 6 years of chronic DR-HC replacement, BMD and bone markers remained stable. TBS decline is more likely due to an age-related derangement of bone microarchitecture rather than a glucocorticoid effect. Our data confirm the safety of DR-HC replacement on bone health in both PAI and SAI patients.

SAT277 Intracellular Cortisol Control: An Approach To Reduce Cushing’s Syndrome Morbidity With Minimal Adrenal Insufficiency Risk?

Abstract Disclosure: F.S. Czerwiec: Employee; Self; Sparrow Pharmaceuticals, Inc. Stock Owner; Self; Sparrow Pharmaceuticals, Inc. D.A. Katz: Employee; Self; Sparrow Pharmaceuticals, Inc. Stock Owner; Self; Sparrow Pharmaceuticals, Inc. P.M. Stewart: Advisory Board Member; Self; Sparrow Pharmaceuticals, Inc. Consulting Fee; Self; Sparrow Pharmaceuticals, Inc. Endocrinologists focus on circulating and excreted cortisol to diagnose and to assess severity and treatment response in Cushing’s syndrome (Cs), but tissue action that drives morbidity is mediated by cortisol binding to intracellular glucocorticoid (GC), mineralocorticoid, and non-genomic receptors. Patients with severe hypercortisolism and deficiency in 11β-hydroxysteroid dehydrogenase type 1 (HSD-1) (an enzyme that generates active GC in target tissues) had no cortisol-related morbidity. Mice without the Hsd11b1 gene (which encodes HSD-1) or given a HSD-1 inhibitor were protected from glucose intolerance, hyperinsulinemia, hepatic steatosis, adiposity, hypertension, myopathy, dermal atrophy, and trabecular bone loss associated with GC administration. The HSD-1 inhibitor AZD4017 prevented detrimental prednisolone effects on glycemic control and osteocalcin in a Phase 1 clinical trial. Although patients with cortisone reductase (i.e., HSD-1) deficiency show no signs or symptoms of adrenal insufficiency (AI), the theoretical possibility of AI in HSD-1 inhibitor treated patients has been raised. Healthy adults received the HSD-1 inhibitor SPI-62 or placebo for 14 days. Data after last dose were analyzed by ANCOVA with baseline as covariate. SPI-62 reduced 24h urinary cortisol metabolites (4.6 +/- 0.3 v 8.0 +/- 0.5 nmol), indicative of a 42% intrahepatocellular cortisol decrease. Results from other inhibitors support HSD-1 as the source of nearly half of intrahepatocellular cortisol in human. Morning serum (293.9 +/- 18.9 v 273.4 +/- 39.3 nM) and 24h urinary (100.7 +/- 6.9 v 89.6 +/- 14.0 nmol) cortisol did not differ between SPI-62 and placebo. No pattern of adverse events suggestive of AI has been observed in SPI-62, nor other HSD-1 inhibitor, clinical trials. Administration of a HSD-1 inhibitor to patients with GC excess could reduce Cs morbidity via reduction of cortisol or GC medication (e.g., prednisolone) available to intracellular receptors. Unlike current Cs treatments that are all associated with substantial AI risk, under HSD-1 inhibition ACTH-mediated cortisol secretion maintains normal circulatory cortisol available to enter cells and thus prevent AI. Data suggest that AI will not be an issue in patients treated with HSD-1 inhibitors. That prediction is under evaluation in Phase 2 clinical trials of SPI-62 in patients with ACTH-dependent Cs, autonomous cortisol secretion, and (together with prednisolone) polymyalgia rheumatica. Until more experience is gathered, caution in patients with modest autonomous cortisol secretion, suppressed ACTH, and atrophied adrenal tissue is advised. In situations that require GC rescue dosing during HSD-1 inhibitor administration, prodrug GCs (i.e., cortisone acetate, prednisone) will not be effective a. s they require HSD-1 for activation. Those patients require active agents (e.g., hydrocortisone, prednisolone). Presentation: Saturday, June 17, 2023

SAT296 Ambulatory 24-hour Tissue Microdialysis Of Cortisol And Metabolites, A Tool For Precision Management In Addison’s Disease

Abstract Disclosure: I. Botusan: None. K. Berinder: None. P. Methlie: None. T. Upton: None. E. Zavala: None. K. Simunkova: None. D.A. Vassiliadi: None. I. Marinelli: None. G.M. Russell: None. M.A. Grytaas: None. S. Tsagarakis: None. S.L. Lightman: None. E.S. Husebye: None. O. Kampe: None. M. Oksnes: None. S. Bensing: None. Background: The autoimmune attack on the adrenals in AD (Addison’s disease) results in lack of the vital hormones cortisol and aldosterone, rendering AD patients dependent on life-long hormone replacement therapy. Current therapies cannot replicate the physiological well-tuned, circadian and ultradian hormonal oscillations properly, probably contributing to increased mortality, morbidity and reduced quality of life. No established biomarker for treatment monitoring exists hampering treatment optimization. Ambulatory 24-hour dynamic steroid profiling may represent a novel therapy-monitoring tool, providing new insights in different replacement therapy regimes. Aim: We aimed to assess 24-hour profiles of free cortisol and metabolites in patients with AD on different glucocorticoid replacement therapies (GRT) and compare the results with healthy volunteers (HV). Material and methods: The study includes 41 patients with AD on different GRT regimes (hydrocortisone, cortisone acetate, modified release hydrocortisone) and 214 healthy participants. Twenty minute microdialysis fractions were collected ambulatory from abdominal subcutaneous tissue over 24 hours using a novel and portable microdialysis fraction collector (U-RHYTHM). Steroid hormones were analyzed by ultrasensitive liquid chromatography tandem mass spectroscopy (LC-MS/MS). Dynamic biomarkers were used to define hormone parameters in both healthy and hormone-replaced participants, and to assess the “time in range” for patients on GRT compared with the profiles of healthy controls. Results: Free cortisol and cortisone levels were measurable in all AD patients, while 18- hydroxycortisol, and corticosterone were detected only in a few AD patients suggesting the persistence of residual adrenal function. Lower cortisol levels during night were observed in AD compared with controls. Ambulatory 24-hour dynamic steroid profiling revealed marked interindividual variations partially depending on the type of GRT and dosing-regime. Conclusions: The 24-hour profiles of free cortisol and cortisone differs between patients with AD and healthy volunteers. Ambulatory 24-hour microdialysis samplings may provide a novel dynamic tool to monitor GRT in AD providing opportunity for individual dose adjustment. Presentation: Saturday, June 17, 2023

Standardised flavonoid extract from Diospyros kaki L.f leaves alleviates high blood pressure and ventricular hypertrophy on cortisone acetate-induced hypertensive rats

The current study was designed to investigate the anti-hypertensive effects of a standardised flavonoid extract from Diospyros kaki L.f leaves (DK extract) on cortisone acetate-induced hypertensive rats. Hypertension was induced in Wistar rats by subcutaneously injecting 2.5 mg cortisone acetate daily and by maintaining the rats on a 1% sodium chloride diet for a period of 28 days. The hypertensive rats were treated daily with DK extract (50 and 100 mg/kg of body weight; p.o.) or captopril (20 mg/kg of body weight; p.o.), a reference drug, for 14 consecutive days. The blood pressure and heart rate of awake rats were measured using a non-invasive tail-cuff method. The heart weight and left ventricular wall thickness of rats were measured. In vitro angiotensin-converting enzyme (ACE) inhibition activity of the DK extract and captopril were analysed. DK extract significantly and dose-dependently reduced systolic and diastolic blood pressure in the hypertensive rats. There were no significant changes observed in heart rate. The cortisone acetate-induced hypertensive rats had increased heart weight and left ventricular wall thickness. These actions were attenuated by the treatment of DK extract (100 mg/kg) and captopril. Moreover, DK extract inhibited ACE activity in vitro with an IC50 of 4.71±0.53 μg/ml. Our findings demonstrate that DK extract exerted anti-hypertensive effects and alleviated ventricular hypertrophy in the cortisone acetate-induced hypertensive rats and these actions occurred at least in part via ACE activity inhibition.

Immune checkpoint inhibitor-associated adrenal insufficiency in Chinese cancer patients: a retrospective analysis.

Immune checkpoint inhibitors (ICIs) are of great success in cancer therapy. This study aimed to identify adrenal insufficiency (AI) associated with immune checkpoint inhibitor (ICI) treatment in cancer patients receiving steroid replacement therapy and report the clinical characteristics of ICI-associated AI and concurrent immune-associated adverse events (irAEs). Patients prescribed cortisone acetate between January 2020 and March 2022 were reviewed to identify AI associated with ICI treatment. Data collected included indication of ICI (cancer type), drug characteristics, and outcomes. A total of 101 patients were diagnosed with AI following treatment with ICIs. The median age was 64years (range 22-83years); 73.3% of the patients were male. Median time to develop primary AI and secondary AI after starting ICI therapy was 200.5 (35-280) days and 178 (16-562) days, respectively. Concurrent irAEs occurred in 67 (66.3%) patients and included 63 (62.4%) endocrine irAEs. Log-rank test showed that there was a trend toward higher likelihood of death at 120-day follow-up in patients initially receiving intravenous hydrocortisone compared with those receiving oral cortisone acetate after diagnosis of AI (p = 0.029). This retrospective study comprehensively documented the clinical characterization of ICI-associated AI. Those initially receiving intravenous hydrocortisone after diagnosis of AI were associated with higher likelihood of death. Physicians should be aware of the variability of ICI-associated irAEs early in the treatment, early diagnoses, and timely management should be made.

Development of a UPLC-ESI-MS/MS method for the determination of triamcinolone acetonide in human plasma and evaluation of its bioequivalence after a single intramuscular injection in healthy volunteers.

Introduction: Triamcinolone acetonide (TA) is commonly used in the treatment of various inflammatory conditions. To ensure its efficacy and safety, it is important to accurately determine its concentration in human plasma and evaluate its bioequivalence. In this study, an efficient ultra-performance liquid chromatography-electrospray ionization-tandem mass spectrometry (UPLC-ESI-MS/MS) method was developed for the quantification of TA in human plasma after a single intramuscular injection. The internal standard used in this method was cortisone acetate (CA). Methods: TA and CA were extracted from plasma using ethyl acetate and N-hexane (4:1, v/v), separated on a C18 reverse-phase column with a mobile phase of acetonitrile-water containing 1% formic acid (55:45, v/v), and analyzed by UPLC-ESI-MS/MS. Multiple-reaction monitoring was performed using the transitions m/z 435.4→397.3 for TA and m/z 403.4→163.1 for CA. Results: The developed UPLC-ESI-MS/MS method demonstrated linearity over a concentration range of 0.53-21.20ng/mL, with a lower limit of quantification of 0.53ng/mL. The intra- and inter-run precision values ranged from 3.007% to 9.960% and 3.528% to 11.26%, respectively. The intra- and inter-run accuracy ranges were -1.962% to -6.577% and -3.371% to 0.348%, respectively. The matrix effect, extraction recovery, and stability of TA all met the acceptance criteria recommended by the National Medical Products Administration (NMPA) for bioassays. In healthy volunteers who received a single intramuscular injection of 80mg of either the test or reference formulation of TA, various pharmacokinetic parameters were determined. C max was found to be 8.616 ± 1.232 and 8.285 ± 1.218ng/mL for the test and reference formulations, respectively. T max was approximately 1.833 ± 0.243 and 1.861 ± 0.230h. The t 1/2 was calculated to be 181.249 ± 78.585 and 201.782 ± 83.551h. AUC 0-720 was 835.642 ± 297.209 and 830.684 ± 331.168ngh/mL, AUC 0-∞ was 991.859 ± 355.939 and 1018.665 ± 420.769ngh/mL for the test and reference formulations, respectively. The average relative bioavailability of TA, determined using AUC 0-720, was 105.4 ± 26.9%. Bioequivalence was evaluated through variance analysis and a double unilateral test, and the 90% confidence intervals of AUC 0-720, C max, and AUC 0-∞ were 92.8%-113.4%, 99.1%-109.1%, and 89.7%-110.9%, respectively (all p > 0.05). Discussion: These results met the bioequivalence criteria set by the NMPA, indicating that the developed UPLC-ESI-MS/MS method accurately determined TA concentrations in the plasma of healthy Chinese volunteers and that the test and reference formulations exhibited bioequivalence in these individuals.

Tissue Glucocorticoid Metabolism in Adrenal Insufficiency: A Prospective Study of Dual-release Hydrocortisone Therapy.

Patients with adrenal insufficiency (AI) require life-long glucocorticoid (GC) replacement therapy. Within tissues, cortisol (F) availability is under the control of the isozymes of 11β-hydroxysteroid dehydrogenase (11β-HSD). We hypothesize that corticosteroid metabolism is altered in patients with AI because of the nonphysiological pattern of current immediate release hydrocortisone (IR-HC) replacement therapy. The use of a once-daily dual-release hydrocortisone (DR-HC) preparation, (Plenadren®), offers a more physiological cortisol profile and may alter corticosteroid metabolism in vivo. Prospective crossover study assessing the impact of 12 weeks of DR-HC on systemic GC metabolism (urinary steroid metabolome profiling), cortisol activation in the liver (cortisone acetate challenge test), and subcutaneous adipose tissue (microdialysis, biopsy for gene expression analysis) in 51 patients with AI (primary and secondary) in comparison to IR-HC treatment and age- and BMI-matched controls. Patients with AI receiving IR-HC had a higher median 24-hour urinary excretion of cortisol compared with healthy controls (72.1 µg/24 hours [IQR 43.6-124.2] vs 51.9 µg/24 hours [35.5-72.3], P = .02), with lower global activity of 11β-HSD2 and higher 5-alpha reductase activity. Following the switch from IR-HC to DR-HC therapy, there was a significant reduction in urinary cortisol and total GC metabolite excretion, which was most significant in the evening. There was an increase in 11β-HSD2 activity. Hepatic 11β-HSD1 activity was not significantly altered after switching to DR-HC, but there was a significant reduction in the expression and activity of 11β-HSD1 in subcutaneous adipose tissue. Using comprehensive in vivo techniques, we have demonstrated abnormalities in corticosteroid metabolism in patients with primary and secondary AI receiving IR-HC. This dysregulation of pre-receptor glucocorticoid metabolism results in enhanced glucocorticoid activation in adipose tissue, which was ameliorated by treatment with DR-HC.

Age-associated features of norovirus infection analysed in mice.

Norovirus (NoV) is the leading global cause of viral gastroenteritis. Young children bear the highest burden of disease and play a key role in viral transmission throughout the population. However, which host factors contribute to age-associated variability in NoV severity and shedding are not well-defined. The murine NoV (MNoV) strain CR6 causes persistent infection in adult mice and targets intestinal tuft cells. Here we find that natural transmission of CR6 from infected dams occurred only in juvenile mice. Direct oral CR6 inoculation of wild-type neonatal mice led to accumulation of viral RNA in the ileum and prolonged shedding in the stool that was replication-independent. This viral exposure induced both innate and adaptive immune responses including interferon-stimulated gene expression and MNoV-specific antibody responses. Interestingly, viral uptake depended on passive ileal absorption of luminal virus, a process blocked by cortisone acetate administration, which prevented ileal viral RNA accumulation. Neonates lacking interferon signalling in haematopoietic cells were susceptible to productive infection, viral dissemination and lethality, which depended on the canonical MNoV receptor CD300LF. Together, our findings reveal developmentally associated aspects of persistent MNoV infection, including distinct tissue and cellular tropism, mechanisms of interferon regulation and severity of infection in the absence of interferon signalling. These emphasize the importance of defining viral pathogenesis phenotypes across the developmental spectrum and highlight passive viral uptake as an important contributor to enteric infections in early life.

Analysis of the effect of triclosan on gonadal differentiation of zebrafish based on metabolome

Although the previous research confirmed that triclosan (TCS) affects the female proportion at the early stage of zebrafish (Danio rerio) and has an estrogen effect, the mechanism by which TCS affects the sex differentiation of zebrafish is not entirely clear. In this study, zebrafish embryos were exposed to different concentrations of TCS (0, 2, 10, and 50 μg/L) for 50 consecutive days. The expression of sex differentiation related genes and metabolites were then determined in larvae using reverse transcription quantitative polymerase chain reaction (RT-qPCR) and Liquid Chromatography-Mass Spectrometer (LC-MS), respectively. TCS upregulated the expression of the sox9a, dmrt1a and amh genes, down-regulating the expression of wnt4a, cyp19a1b, cyp19a1a, and vtg2 gene. The overlapped classification of Significant Differential Metabolites (SDMs) between the control group and three TCS treated groups related to gonadal differentiation was Steroids and steroid derivatives, including 24 down-regulated SDMs. The enriched pathways related to gonadal differentiation were Steroid hormone biosynthesis, Retinol metabolism, Metabolism of xenobiotics by cytochrome P450, and Cortisol synthesis and secretion. Moreover, SDMs were significantly enriched in Steroid hormone biosynthesis in the 2 μg/L TCS group, which included Dihydrotestosterone, Cortisol, 11beta-hydroxyandrost-4-ene-3, 17-dione, 21-Hydroxypregnenolone, Androsterone, Androsterone glucuronide, Estriol, Estradiol, 19-Hydroxytestosterone, Cholesterol, Testosterone, and Cortisone acetate. Results showed that TCS affects the female proportion mainly through Steroid hormone biosynthesis, in which aromatase plays a key role in zebrafish. Retinol metabolism, metabolism of xenobiotics by cytochrome P450, and cortisol synthesis and secretion may also participate in TCS-mediated sex differentiation. These findings reveal the molecular mechanisms of TCS-induced sex differentiation, and provide theoretical guidance for the maintenance of water ecological balance.

Analysis of interaction effects of persulphate and peroxymonosulphate on solar photocatalytic degradation of cortisone acetate

AbstractThe degradation of environmental contaminants using sustainable solar energy is one of the most promising applications of the photocatalytic process. In this research, the individual and interaction effects of persulphate (PS) and peroxymonosulphate (PMS) oxidants on the solar photocatalytic degradation of cortisone acetate (CA) were studied. Concentrations of PS, PMS, and photocatalyst were independent variables, and the response was CA degradation efficiency. Response surface methodology (RSM) and artificial neural network (ANN) models were developed to predict the CA degradation efficiency. The analysis of the current results revealed that the optimum amounts of individual variables were highly affected by the other variables, which confirmed their significant interaction effect. As an example, by increasing the PS concentration, not only were the required concentrations of PMS and photocatalyst decreased, but also the degradation efficiency was enhanced. Then, the overall optimum concentration of the photocatalyst, PS, and PMS were found to be respectively 328.7, 119.1, and 194.2 mg/L using the genetic algorithm method. The maximum CA degradation efficiency at the optimum condition was 95.6% after only 30 min of solar radiation. Finally, investigation of relative importance of the variables showed that the concentrations of both oxidants affected the degradation efficiency almost equally.

ITRAQ-Based Quantitative Proteomic Analysis of Arthrobacter simplex in Response to Cortisone Acetate and Its Mutants with Improved Δ1-Dehydrogenation Efficiency.

Arthrobacter simplex is extensively used for cortisone acetate (CA) biotransformation in industry, but the Δ1-dehydrogenation molecular fundamental remains unclear. Herein, the comparative proteome revealed several proteins with the potential role in this reaction, which were mainly involved in lipid or amino acid transport and metabolism, energy production and conversion, steroid degradation, and transporter. The influences of six proteins were further confirmed, where pps, MceGA, yrbE4AA, yrbE4BA, and hyp2 showed positive impacts, while hyp1 exhibited a negative effect. Additionally, KsdD5 behaved as the best catalytic enzyme. By the combined manipulation in multiple genes under the control of a stronger promoter, an optimal strain with better catalytic enzyme activity, substrate transportation, and cell stress tolerance was created. After biotechnology optimization, the production peak and productivity were, respectively, boosted by 4.1- and 4.0-fold relative to the initial level. Our work broadens the understanding of the Δ1-dehydrogenation mechanism, also providing effective strategies for excellent steroid-transforming strains.

Effects of the therapy shift from cortisone acetate to modified-release hydrocortisone in a group of patients with adrenal insufficiency.

Patients with adrenal insufficiency (AI) may be exposed to supraphysiological glucocorticoids levels during standard treatment with cortisone acetate (CA) or immediate-release hydrocortisone (IR-HC). Recent studies, predominantly including patients in IR-HC treatment, suggested that modified-release hydrocortisone (MRH) provide a more physiological cortisol rhythm, improving metabolic control and quality of life. Our primary aim was to assess clinical and biochemical modifications in patients shifted from CA to MRH. We designed a retrospective longitudinal study, enrolling 45 AI patients (22 primary and 23 secondary AI) treated exclusively with CA thrice daily, shifted to MRH once daily; 29/45 patients concluded at least 18-months follow-up (MRH-group). We recruited 35 AI patients continuing CA as a control group (CA-group). Biochemical and clinical data, including metabolic parameters, bone quality, and symptoms of under- or overtreatment were collected. In 24 patients, a daily salivary cortisol curve (SCC) performed before and one month after shifting to MRH was compared to healthy subjects (HS). No significant changes in glycometabolic and bone parameters were observed both in MRH and CA-groups during a median follow-up of 35 months. A more frequent decrease in blood pressure values (23.1% vs 2.8%, p=0.04) and improvement of under- or overtreatment symptoms were observed in MRH vs CA-group. The SCC showed a significant steroid overexposure in both CA and MRH-groups compared to HS [AUC (area under the curve) = 74.4 ± 38.1 nmol×hr/L and 94.6 ± 62.5 nmol×hr/L respectively, vs 44.1 ± 8.4 nmol×hr/L, p<0.01 for both comparisons], although SCC profile was more similar to HS in MRH-group. In our experience, patients shifted from CA to equivalent doses of MRH do not show significant glycometabolic modifications but blood pressure control and symptoms of over-or undertreatment may improve. The lack of amelioration in glucose metabolism and total cortisol daily exposure could suggest the need for a dose reduction when shifting from CA to MRH, due to their different pharmacokinetics.

Development of a Corticosteroid-Immunosuppressed Mouse Model to Study the Pathogenesis and Therapy of Influenza-Associated Pulmonary Aspergillosis.

Influenza-associated pulmonary aspergillosis (IAPA) is a feared complication in patients with influenza tracheobronchitis, especially those receiving corticosteroids. Herein, we established a novel IAPA mouse model with low-inoculum Aspergillus infection and compared outcomes in mice with and without cortisone acetate (CA) immunosuppression. CA was an independent predictor of increased morbidity/mortality in mice with IAPA. Early antifungal treatment with liposomal amphotericin B was pivotal to improve IAPA outcomes in CA-immunosuppressed mice, even after prior antiviral therapy with oseltamivir. In summary, our model recapitulates key clinical features of IAPA and provides a robust preclinical platform to study the pathogenesis and treatment of IAPA.

97. A Cross-kingdom Vaccine Protects against Multiple Healthcare-associated Infections

Abstract Background Rise in multi-drug-resistance (MDR) among pathogens and increasing populations at risk for these infections raises the threat of nearly untreatable infectious diseases. Thus, novel vaccine strategies to prevent and/or treat MDR pathogens would benefit global health immensely. Candida albicans (CA) cell wall proteins Als3p and Hyr1p protect against candidemia due to CA and MDR C. auris (CAU). Further, Hyr1p shares structural homology with conserved hemagglutinin (FhaB) and OmpA proteins in many Gram-negative bacteria (GNB) Acinetobacter baumannii (AB), Klebsiella pneumoniae (KP), and Pseudomonas aeruginosa (PA). Thus, Hyr1p antigen-based active and passive vaccinations protect against AB and KP pneumonia. We hypothesized that a dual Als3p/Hyr1p antigen vaccine formulated with CAF01 (a clinical stage adjuvant) that can promote a balanced Th1/Th2/Th17 immune response is likely to protect against multiple healthcare-associated infections caused by Candida species and MDR GNB. Methods To formulate Als3p/Hyr1p antigen, we mixed CAF01 with Als3p and Hyr1p antigens in the following ratios: 0/0, 10/10, 10/30, 30/10 and 30/3 µg for each dose. CD-1 mice with each dose subcutaneously, followed by booster immunization on day 21. For efficacy evaluation, vaccinated mice (n=20 mice/group) were immunosuppressed by administration of cyclophosphamide and cortisone acetate on day -2 and +3, relative to infection with either CAU, AB, KP or PA. Infection occurred two weeks following the booster immunization. For, CA infection, immunocompetent mice were infected intravenously two weeks following a primary booster or after a second booster given on day 35. Results All Als3p/Hyr1p formulations induced robust immunity against both antigens. For CA infection, two booster immunizations provide superior protection over one booster immunization. For CAU and GNB, one booster dose was enough to provide significant protection. Further, both 30/10 and 10/10 vaccine formulations protected significantly against all five infections. Specifically, for fungal infections, 30/10 and 10/10 formulations showed 30-40% and 40-50% survival efficacies (vs. 0% for placebo), respectively. TABLE 1. Survival efficacy of Als3p/Hyr1p formulations against Candida species and Gram-negative bacterial infections. Conclusion Our study shows that the Als3p/Hyr1p induced a robust protective immunity against CA, MDR CAU and GNB. Disclosures All Authors: No reported disclosures.

LB1580. Efficacy Assessment of MAT2203 Encochleated Oral Formulation of Amphotericin B, in the Neutropenic Mouse Model of Pulmonary Mucormycosis

Abstract Background Invasive mucormycosis (IM) is associated with high mortality and morbidity. MAT2203 is an encochleated oral formulation of amphotericin B which has been shown to be safe and effective against murine aspergillosis and murine cryptococcal meningoencephalitis. We sought to compare the efficacy of MAT2203 to liposomal amphotericin B (LAMB) treatment in a neutropenic mouse model of IM. Methods ICR mice were immunosuppressed with cyclophosphamide and cortisone acetate on days -2, -3 and +8, relative to infection with intratracheally instilled Rhizopus delemar 99-880 or M. circinelloides f. jenssenii DI15-131. Treatment with placebo (diluent control), oral MAT2203 (5 to 45 mg/kg, given qd or bid for 7 days) or LAMB (10 mg/kg, iv, qd for 4 days), began 16 h post infection. Survival (n=10-20/group from 1/2 experiments) through Day +21 and tissue fungal burden of lungs or brain (n=10/group) euthanized on Day +4 post infection (conidial equivalents using qPCR) served as a primary and secondary endpoint, respectively. Results For Rhizopus delemar infection, doses of MAT2203 5,15 mg/kg qd or 7.5 mg/kg bid, significantly prolonged median survival time (MST) and enhanced overall survival vs. placebo-treated mice (MST of 9 Days and 0% survival for placebo-treated mice vs. 13 Days, and 20-40% for MAT2203 doses, P&amp;lt; 0.05 by Log-Rank). Importantly, MAT2203 treatments were as effective as LAMB (MST of 16 days and overall survival of 45%). For mice infected with M. circinelloides, MAT2203 at 15 mg/kg, qd significantly prolonged MST and enhanced overall survival vs. placebo-treated mice (MST of 5 Days and 0% survival, for placebo-treated mice vs. 13.5 Days and 50% survival for MAT2203, P&amp;lt; 0.05). In both infection models MAT2203 treatment of 15 mg/kg or LAMB resulted in significant ∼1.0-1.5 log reduction and ∼2.0-2.2 log reduction in lung and brain fungal burden vs. placebo, respectively (Wilcoxon Rank Sum). Conclusion MAT2203 demonstrated in vivo efficacy in treating R. delemar or M. circinelloides pulmonary infection in immunosuppressed mice, which was equivalent to the LAMB current standard of care. Continued investigation and development of MAT2203 as a novel, and oral formulation of amphotericin antifungal agent against mucormycosis is warranted. Disclosures Theresa Matkovits, PhD, Matinas BioPharma: Employee Jenel Cobb, PhD, Matinas BioPharma: Employee Raphael J. Mannino, PhD, Matinas BioPharma: Employee Ashraf S. Ibrahim, PhD, Matinas BioPharma: Advisor/Consultant|Matinas BioPharma: Grant/Research Support|SFunga: Grant/Research Support.

499. Immunotherapy with Pattern Recognition Receptor Agonists Improves Morbidity and Mortality in a Corticosteroid-Immunosuppressed Murine Model of Influenza-Associated Pulmonary Aspergillosis

Abstract Background Corticosteroids are known to increase the incidence and worsen the outcomes of influenza-associated pulmonary aspergillosis (IAPA) in patients with severe influenza pneumonia. Adjunct immunomodulatory strategies might improve the treatment of this emerging entity, but in-vivo data in validated corticosteroid-immunosuppressed animal models of IAPA is scarce. Methods 8-10-week-old female BALB/c mice were infected with 7.5% LD90 of a mouse-adapted influenza A/H3N2/Hong Kong/1968 strain, delivered by aerosolization on day (d) 0. On d5 and d8, mice received two intraperitoneal injections of 10 mg cortisone acetate (CA) or mock injections. On d9, mice were intranasally challenged with 50,000 Aspergillus fumigatus (AF) conidia. Infection severity was scored using the viral pneumonia score (VPS, 0 = healthy to 12 = moribund) and the murine sepsis score (MSS, 0 = healthy to 3 = moribund). For therapeutic studies, mice received a 30-minute nebulization of the toll-like receptor (TLR) 2/6/9 agonists Pam2+ODN (PMID: 34153197; 4 µM/1 µM) or PBS (mock treatment) on d8 (single-dose) or d8+12 (dual-dose). Therapeutic studies used a combined morbidity/mortality endpoint, with an event defined as either death or reaching a VPS ≥ 7. Results CA-immunosuppressed mice with IAPA had 64% 16-day mortality and survivors had severe morbidity (median MSS 2.7). In contrast, all non-immunosuppressed mice with IAPA survived and showed consistently low morbidity (median d16 MSS 0.5). All mock-treated CA-immunosuppressed mice with IAPA reached the combined morbidity/mortality endpoint by d11. Single-dose Pam2+ODN treatment led to universal event-free survival until d13 but all mice deteriorated by d16. Dual-dose Pam2+ODN treatment before and after AF infection led to 75% event-free survival until d21, with all survivors fully recovering (VPS = 0, p &amp;lt; 0.001, Figure 1). Figure 1 Conclusion Aligned with clinical evidence, we found detrimental outcomes of IAPA in CA-immunosuppressed mice. Treatment with TLR agonists significantly improved morbidity and mortality of CA-immunosuppressed mice with IAPA. Next, we will study the impact of synergistic anti-infective and immunotherapeutic interventions on the course and immunopathogenesis of IAPA. Disclosures Dimiitrios P. Kontoyiannis, MD, ScD, PhD (hon), AbbVie: Advisor/Consultant|Astellas Pharma: Advisor/Consultant|Astellas Pharma: Grant/Research Support|Astellas Pharma: Honoraria|Cidara Therapeutics: Advisor/Consultant|Gilead Sciences: Advisor/Consultant|Gilead Sciences: Grant/Research Support|Gilead Sciences: Honoraria|Merck: Advisor/Consultant.