Single Iv Research Articles

Comparison of ultrafiltration and microdialysis for ceftriaxone protein-binding determination.

High protein binding (PB) of antibiotics has an impact on their antimicrobial activity. It has been questioned whether in vitro PB determination can capture the dynamic and concentration-dependent PB of highly bound antibiotics. This clinical study compared in vitro ultrafiltration (UF) and in vivo IV microdialysis (MD) methods to determine ceftriaxone PB. Six healthy male volunteers received a single IV 2 g dose of ceftriaxone. Unbound ceftriaxone plasma concentrations were measured with MD and venous plasma sampling with subsequent UF. Pharmacokinetic parameters were determined using non-compartmental pharmacokinetic analysis. Non-linear mixed-effects modelling was used to quantify the PB. The PTA was estimated. The Cmax of ceftriaxone total plasma concentration (297.42 ± 21.0 mg/L) was approximately 5.5-fold higher than for free concentrations obtained with UF (52.83 ± 5.07 mg/L), and only 3.5-fold higher than for free concentrations obtained with MD (81.37 ± 26.93 mg/L). Non-linear, saturable PB binding was confirmed for both UF and MD. Significantly different dissociation constants (Kd) for the albumin/ceftriaxone complex were quantified: in UF it was 23.7 mg/L (95% CI 21.3-26.2) versus 15.9 mg/L (95% CI 13.6-18.6) in MD. Moreover, the estimated number of binding sites (95% CI) per albumin molecule was 0.916 (0.86-0.97) in UF versus 0.548 in MD (0.51-0.59). The PTA obtained with MD was at most 27% higher than with UF. In vitro UF versus in vivo intravasal MD revealed significantly different PB, especially during the distribution phase. The method of PB determination could have an impact on the breakpoint determination and dose optimisation of antibiotics.

Effects of climate variables on the White Stork (Ciconia ciconia L.) productivity in a long term study

Abstract We analysed the effects of weather and climatic patterns on the productivity of the White Stork in Hungary between 1958 and 2017, using i) linear mixed effect models (LMM), ii) LMM-s extended by a single random effect variable or a nested combination; iii) LMM-s extended by a single fixed effect variable and iv) using an additive model of the selected variables. As a preselection, the following climatic variables were identified with substantial support: March mean temperature, March precipitation, April mean temperature, June mean temperature, June precipitation (negative), July mean temperature. The slight increase of the mean number of fledged chicks over 59 years could be the result of the increasing mean temperature, but in itself it might not be strong enough to prove that climate change will overall benefit White Stork productivity. Higher temperature and precipitation values are favourable, probably because of the higher biomass, providing more prey, but high precipitation is unfavourable until the thermoregulation of chicks is not developed. Decreasing amounts of precipitation may cause loss of wetlands as suitable feeding sites. Extreme weather is important to complement the picture given by climate change.

A simple APACHE IV risk dynamic nomogram that incorporates early admitted lactate for the initial assessment of 28-day mortality in critically ill patients with acute myocardial infarction

BackgroundEarly risk stratification is important for patients with acute myocardial infarction (AMI). We aimed to develop a simple APACHE IV dynamic nomogram, combined with easily available clinical parameters within 24 h of admission, thus improving its predictive power to assess the risk of mortality at 28 days.MethodsClinical information on AMI patients was extracted from the eICU database v2.0. A preliminary XGBoost examination of the degree of association between all variables in the database and 28-day mortality was conducted. Univariate and multivariate logistic regression analysis were used to perform screening of variables. Based on the multifactorial analysis, a dynamic nomogram predicting 28-day mortality in these patients was developed. To cope with missing data in records with missing variables, we applied the multiple imputation method. Predictive models are evaluated in three main areas, namely discrimination, calibration, and clinical validity. The discrimination is mainly represented by the area under the receiver operating characteristic curve (AUC), net reclassification improvement (NRI) and integrated discrimination improvement (IDI). Calibration is represented by the calibration plot. Clinical validity is represented by the decision curve analysis (DCA) curve.ResultsA total of 504 people were included in the study. All 504 people were used to build the predictive model, and the internal validation model used a 500-bootstrap method. Multivariate analysis showed that four variables, APACHE IV, the first sample of admission lactate, prior atrial fibrillation (AF), and gender, were included in the nomogram as independent predictors of 28-day mortality in AMI. The prediction model had an AUC of 0.819 (95%CI 0.770–0.868) whereas the internal validation model had an AUC of 0.814 (95%CI 0.765–0.860). Calibration and DCA curves indicated that the dynamic nomogram in this study were reflective of real-world conditions and could be applied clinically. The predictive model composed of these four variables outperformed a single APACHE IV in terms of NRI and IDI. The NRI was 16.4% (95% CI: 6.1–26.8%; p = 0.0019) and the IDI was 16.4% (95% CI: 6.0–26.8%; p = 0.0020). Lactate accounted for nearly half of the total NRI, which showed that lactate was the most important of the other three variables.ConclusionThe prediction model constructed by APACHE IV in combination with the first sample of admission lactate, prior AF, and gender outperformed the APACHE IV scoring system alone in predicting 28-day mortality in AMI. The prediction dynamic nomogram model was published via a website app, allowing clinicians to improve the predictive efficacy of the APACHE IV score by 16.4% in less than 1 min.

Abstract 13097: Safety, Pharmacokinetics, and Pharmacodynamics of AZD3366 Alone and in Combination With Aspirin and Ticagrelor in Healthy Subjects

Introduction: Despite reperfusion in patients with ST-elevation myocardial infarction and acute ischemic stroke; reperfusion injury, microvascular dysfunction, thrombosis, and obstruction remain important causes of persistent ischemia and infarct expansion leading to poor clinical outcome. AZD3366, a recombinant human apyrase engineered to enhance hydrolysis of extracellular adenosine tri-/diphosphate (ATP/ADP) to adenosine monophosphate and subsequently adenosine, has exhibited anti-thrombotic, anti-inflammatory, and tissue protective properties in preclinical studies. Methods: In this phase 1 randomized single-blind placebo-controlled study in healthy subjects ( NCT04588727 ), 7 cohorts of 8 subjects received a single short IV infusion of AZD3366 (2-640 mg [n=6]) or placebo (n=2). A further 3 cohorts of 5 healthy Japanese/1 cohort of 8 healthy Chinese subjects were included within this dose range. A separate group of 24 subjects received 160 mg of AZD3366 in combination with aspirin and ticagrelor (n=12) or aspirin and ticagrelor alone (n=12). The primary objective was to assess safety and tolerability; secondary objectives included assessment of pharmacokinetics (PK) and pharmacodynamics of AZD3366. Results: Overall, 103 subjects were randomized; mean age 37.2 (± 9) years, 100 were male. AZD3366 treatment alone or in combination with ticagrelor and aspirin did not result in any clinically relevant safety or tolerability findings. Peak AZD3366 plasma concentrations were reached within 0.5 h post-dosing, followed by monophasic or biphasic decline with a terminal PK half-life of approximately 140 h. Complete ADP-stimulated platelet aggregation inhibition was reached within 10 mins post-dosing; duration ranged from 4 h at 2 mg to ~35 days at 640 mg (Figure). Conclusions: AZD3366 alone or in combination with aspirin and ticagrelor was generally safe, well tolerated, and achieved complete platelet inhibition with a dose-dependent duration.

Single oral or intravenous administration of voriconazole achieved recommended therapeutic minimum inhibitory concentrations against Aspergillus in the common raven (Corvus corax).

To determine the pharmacokinetics of voriconazole after single IV or orally administered boluses in common ravens (Corvus corax). 8 healthy common ravens. Voriconazole (5 mg/mL, 10 mg/kg IV) was administered to 8 birds, and then plasma voriconazole concentrations were measured at various time points by high-pressure liquid chromatography with mass spectrometry. Starting 6 months later in a randomized 3-treatment 3-period regimen, birds received a single oral dose of voriconazole suspension (10 mg/mL; 6, 12, and 24 mg/kg PO). The study period was May 2015 to March 2016. Voriconazole (10 mg/kg IV) achieved an initial plasma concentration of 6.31 µg/mL when measured over 21 hours. After oral administration of voriconazole at 6, 12, and 24 mg/kg, the relative bioavailability was 67.5%, 209%, and 183%, respectively. For the 6-mg/kg dose, the maximum plasma concentration was reached at 30 minutes after administration and remained in the therapeutic range of 0.5 to 1 µg/mL for approximately 15 hours. The 12- and 24-mg/kg doses resulted in concentrations in a potentially toxic range. Voriconazole was well tolerated. All 4 doses resulted in plasma concentrations of voriconazole > 0.5 µg/mL, which is the minimum inhibitory concentration recommended for pathogenic species of Aspergillus fungi known to affect birds. A single dose of voriconazole administered as 10 mg/kg IV or 6 mg/kg PO resulted in recommended target plasma concentrations. Administration of voriconazole 6 mg/kg PO 2 to 3 times daily may be adequate for treatment without exceeding the toxic range.

Study on pharmacokinetics and tissue distribution of Polygonatum sibiricum polysaccharide in rats by fluorescence labeling

To study the pharmacokinetics and tissue distribution characteristics of Polygonatum sibiricum (P. sibiricum) polysaccharide administered orally and intravenously in rats, the latest quantitative analysis method was established where P. sibiricum polysaccharide was labeled with fluorescein isothiocyanate (FITC) in plasma and tissues. Quantitative analysis method of P. sibiricum polysaccharide in rat plasma and tissues was established by fluorescence spectrophotometry with FITC as a highly sensitive fluorescent molecular probe. The results showed that P. sibiricum polysaccharide was successfully labeled with FITC, and the degree of substitution was 0.55 %. Pharmacokinetic characteristics showed that oral administration (ig) and intravenous injection (iv) were consistent with the characteristics of two-compartment model. PRP-TYR-FITC administered orally was poorly absorbed in rats with low bioavailability. After a single ig and iv administration in rats for 8 h, P. sibiricum polysaccharide can be distributed in most tissues. The analysis results showed that P. sibiricum polysaccharide was distributed mostly in lung, kidney and liver for both routes of administration. When taking orally, the distribution pattern was: lung > liver > kidney > small intestine > stomach > heart > spleen > brain. When taking intravenously, the distribution pattern was: liver > lung > kidney > small intestine > heart > stomach > spleen > brain. Fluorescence labeling of P. sibiricum polysaccharide by FITC was successfully realized. This method was proved to be suitable for the study of pharmacokinetics and tissue distribution of P. sibiricum polysaccharide in rats. The above research lays foundation for further elucidating the clinical pharmacological mechanism of polysaccharide in P. sibiricum.

Abstract 1765: Discovery and first disclosure of AZD8205, a B7-H4-targeted antibody-drug conjugate utilizing a novel topoisomerase I linker-warhead

Abstract The cell-surface glycoprotein B7-H4 is overexpressed in a range of solid tumors including breast cancer, ovarian serous carcinoma, endometrial carcinoma, and cholangiocarcinoma, yet has limited expression in normal tissue, making it an attractive target for an antibody-drug conjugate (ADC). This presentation describes for the first time the development of AZD8205, a B7-H4 targeted ADC incorporating a novel topoisomerase 1 inhibitor (TOP1i) linker-warhead, AZ’0133 which was designed to exploit the full potential of B7-H4 as an ADC target. Initially, we investigated a series of more than 35 TOP1i compounds as warheads and achieved activity in a clinically relevant nM range. We further optimized the conjugation site and chemistry to reduce the potential for aggregation while maintaining potency, overcoming major synthetic challenges to deliver a robust synthetic route amenable to scale-up. Finally, with a series of optimized linker-warheads, we explored the impact of linker-warhead design on ADC hydrophobicity, stability, efficacy, pharmacokinetics and tolerability culminating in the development of AZD8205. The primary mechanism of action of AZD8205 is intracellular delivery of the TOP1i warhead to B7-H4 positive cells, leading to DNA damage and apoptotic cell death. AZD8205 drove bystander killing of target negative cells in mixed cultures in vitro, which is further supported by robust antitumor activity observed in in vivo studies with patient-derived xenograft (PDX) tumors with heterogeneous target expression, representing multiple tumor indications. In a study of 26 human TNBC PDX tumors, a single IV administration of 3.5 mg/kg AZD8205 provided an overall response rate of 69% (tumor regression of 30% or greater from baseline) and complete responses observed in 9/26 (36%) of models. To understand the biology underlying antitumor response, we conducted a multiparametric analysis including genomics, proteomics and computational pathology and found that deeper antitumor activity was observed in models with elevated B7-H4 expression as well as in models with defects in DNA damage repair (DDR). To further exploit the DNA damage elicited by the TOP1i warhead, we examined combinations of AZD8205 with small molecules, including a novel PARP1 selective inhibitor, in a BRCA wild type MDA-MB-468 model. These data suggest that AZD8205 is a promising therapeutic candidate for the treatment of B7-H4 positive solid tumors. A first in human phase 1 study in patients with advanced solid tumors is currently ongoing (NCT05123482). Citation Format: Krista Kinneer, Niall J. Dickinson, Luke Masterson, Thais Cailleau, Ian Hutchinson, Balakumar Vijayakrishnan, Nazzareno Dimasi, R. James Christie, Mary McFarlane, Kathryn Ball, Arthur Lewis, Sofia Koch, Lee Brown, Yue Huang, Anton I. Rosenbaum, Jiaqi Yuan, Si Mou, Noel R. Monks, Jon Chesebrough, Ravinder Tammali, Judith Anderton, Darrin Sabol, Frances Anne Tosto, Philipp Wortmann, Zachary A. Cooper, Pauline Ryan, John Hood, Carlos Fernandez Teruel, Carlos Serra Traynor, Andy Pike, Michael Davies, Elisabetta Leo, Kimberly Cook, Nadia Luheshi, Philip W. Howard, Puja Sapra. Discovery and first disclosure of AZD8205, a B7-H4-targeted antibody-drug conjugate utilizing a novel topoisomerase I linker-warhead [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1765.

Pharmacokinetics of isavuconazole in healthy cats after oral and intravenous administration.

BackgroundIsavuconazole is a triazole antifungal drug that has shown good efficacy in human patients. Absorption and pharmacokinetics have not been evaluated in cats.ObjectivesTo determine the pharmacokinetics of isavuconazole in cats given a single IV or PO dose.AnimalsEight healthy, adult research cats.MethodsFour cats received 100 mg capsules of isavuconazole PO. Four cats received 5 mg/kg isavuconazole solution IV. Serum was collected at predetermined intervals for analysis using ultra‐high performance liquid chromatography‐tandem mass spectrometry. Data were analyzed using a 2‐compartment uniform weighting pharmacokinetic analysis with lag time for PO administration and a 2 compartment, 1/y2 weighting for IV administration. Predicted 24 and 48‐hour dosing intervals of 100 mg isavuconazole administered PO were modeled and in vitro plasma protein binding was assessed.ResultsBoth PO and IV drug administration resulted in high serum concentrations. Intravenous and PO formulations of isavuconazole appear to be able to be used interchangeably. Peak serum isavuconazole concentrations occurred 5 ± 3.8 hours after PO administration with an elimination rate half‐life of 66.2 ± 55.3 hours. Intersubject variability was apparent in both the PO and IV groups. Two cats vomited 6 to 8 hours after PO administration. No adverse effects were observed in the IV group. Oral bioavailability was estimated to be approximately 88%. Serum protein binding was calculated to be approximately 99.0% ± 0.03%.Conclusions and Clinical ImportanceIsavuconazole might prove to be useful in cats with fungal disease given its favorable pharmacokinetics. Additional studies on safety, efficacy, and tolerability of long‐term isavuconazole use are needed.

Solutions à la pénurie de médecins de famille

The disposition and tissue distribution of angiopeptin, a long-acting octapeptide analogue of somatostatin, were studied in rats following single iv and sc administration of the drug. Similar plasma levels and excretion values of angiopeptin were observed by using radioimmunoassay and radiolabeling techniques. Angiopeptin was absorbed fairly rapidly, with a mean peak plasma level of 25 +/- 4.1 ng/ml at 10-15 min after administration. The kinetics of angiopeptin following sc administration closely resembled those following iv administration due to rapid absorption. The pharmacokinetics of angiopeptin can be described by a two-compartment model. The plasma half-life of the drug ranged from 2.6-2.9 hr when administered sc and 1.98-2.5 hr when given iv. Distribution of angiopeptin was rapid, with the highest concentration appearing in the liver. Half-lives in the liver and bile were short. Most of the drug was excreted in the feces via the bile, while approximately 10% was excreted in the urine. Angiopeptin was also found to be secreted in the saliva. TLC and HPLC of blood, urine, feces, and bile samples did not reveal the presence of any metabolites. In conclusion, the in vivo fate of angiopeptin is characterized by little or no hepatic metabolism and rapid biliary excretion.

Magnetic resonance imaging contrast-enhancement with superparamagnetic iron oxide nanoparticles amplifies macrophage foam cell apoptosis in human and murine atherosclerosis.

(Ultra) Small superparamagnetic iron oxide nanoparticles, (U)SPIO, are widely used as magnetic resonance imaging contrast media and assumed to be safe for clinical applications in cardiovascular disease. As safety tests largely relied on normolipidaemic models, not fully representative of the clinical setting, we investigated the impact of (U)SPIOs on disease-relevant endpoints in hyperlipidaemic models of atherosclerosis. RAW264.7 foam cells, exposed in vitro to ferumoxide (dextran-coated SPIO), ferumoxtran (dextran-coated USPIO), or ferumoxytol [carboxymethyl (CM) dextran-coated USPIO] (all 1 mg Fe/mL) showed increased apoptosis and reactive oxygen species accumulation for ferumoxide and ferumoxtran, whereas ferumoxytol was tolerated well. Pro-apoptotic (TUNEL+) and pro-oxidant activity of ferumoxide (0.3 mg Fe/kg) and ferumoxtran (1 mg Fe/kg) were confirmed in plaque, spleen, and liver of hyperlipidaemic ApoE-/- (n = 9/group) and LDLR-/- (n = 9-16/group) mice that had received single IV injections compared with saline-treated controls. Again, ferumoxytol treatment (1 mg Fe/kg) failed to induce apoptosis or oxidative stress in these tissues. Concomitant antioxidant treatment (EUK-8/EUK-134) largely prevented these effects in vitro (-68%, P < 0.05) and in plaques from LDLR-/- mice (-60%, P < 0.001, n = 8/group). Repeated ferumoxtran injections of LDLR-/- mice with pre-existing atherosclerosis enhanced plaque inflammation and apoptosis but did not alter plaque size. Strikingly, carotid artery plaques of endarterectomy patients who received ferumoxtran (2.6 mg Fe/kg) before surgery (n = 9) also showed five-fold increased apoptosis (18.2 vs. 3.7%, respectively; P = 0.004) compared with controls who did not receive ferumoxtran. Mechanistically, neither coating nor particle size seemed accountable for the observed cytotoxicity of ferumoxide and ferumoxtran. Ferumoxide and ferumoxtran, but not ferumoxytol, induced apoptosis of lipid-laden macrophages in human and murine atherosclerosis, potentially impacting disease progression in patients with advanced atherosclerosis.

Role of Dexamethasone in Post-tonsillectomy Morbidities

Objective: The aim of this study was to find out the effect of Dexamethasone on pain following tonsillectomy. In addition topost-tonsillectomy pain, this study also explore the comparative outcome of use of additional analgesic, fever, nausea &amp;vomiting as well as oral intake of semisolid food between the groups. Methods: A double blind, randomized control trial using a single pre-operative dose of Dexamethasone (0.15mg/kg bodyweight) or normal saline in the randomly selected 100 patients of age more than 3 years, undergoing tonsillectomy fulfillinginclusion criteria of this study, in the Department of Otolaryngology and head Neck surgery, Dhaka Medical College Hospital,Dhaka during July 2009 to June 2010. Surgical techniques, anesthetic drugs, use of analgesic and other medication werestandardized. Both groups were assessed clinically at 6, 12 and 24 hours post-operatively for pain, requirement of analgesic,fever, nausea &amp; vomiting, oral intake, condition of the tonsillar fossa and reactionary haemorrhage. Visual Analogue Scale(VAS) used for the assessment of pain. Results: Post-operative pain was assessed by Visual Analogue Scale (VAS) in both Dexamethasone and Control groups.Mean+/-SD of the VAS in Dexamethasone and Control groups at 6 hours was 5.04 +/- 1.09 vs 5.96 +/- 1.03 (p= &lt; 0.0001), at 12hours was 3.94 +/- 0.93 vs 5.14 +/- 0.61 (p=&lt; 0.0001) and at 24 hours was 2.64 +/- 0.69 vs 3.84 +/- 0.87 (p=&lt; 0.0001). Requirement ofadditional analgesic (p&lt;0.05), fever (P&lt;0.001), Post-Operative Nausea and Vomiting (PONV) (P &lt; 0.005), oral intake ofsemisolid food (P&lt;0.005) between Dexamethasone and control group were significantly different. Regarding condition of thetonsillar fossa and reactionary haemorrhage, no significant differences were observed. Conclusion: A single preoperative IV dose of Dexamethasone (0.15 mg/kg), given just before starting surgery, provided goodand prolonged analgesia. It also reduces fever, nausea &amp; vomiting and odynophagia. Those enhance earlier and betterpostoperative quality of oral intake and decrease morbidities.

Intravenous Dexmedetomidine versus Intravenous Esmolol in Blunting the Laryngoscopy Response in Adult Normotensives Undergoing Elective Surgeries: A Randomised Clinical Trial

Introduction: A wide array of drugs are available for blunting the laryngoscopy response. Amongst them both dexmedetomidine, and esmolol belong to the non-opioid group having least interference with the recovery process without causing significant respiratory depression, thus are suitable intervention for this purpose. Dexmedetomidine a sedative highly selective α2 adrenoceptor agonist and has an anaesthetic-sparing effect at induction. It suppresses the release of catecholamine in response to a noxious stimulant because of its central sympatholytic action. Esmolol with a different pharmacokinetic profile is a water soluble, cardio-selective, an ultrashort acting beta blocker has a short t1/2 thus suited for suppressing the transient pressor reflexes following acute noxious surgical or anesthesia stimuli. Aim: To compare the degree of attenuation of the laryngoscopy response following the use of single preinduction dose (Intravenous infusion at a dose of 1 µg/kg) of dexmedetomidine with that of esmolol (intravenous bolus at a dose of 0.5 mg/kg) in adult normotensives undergoing elective intubations. Materials and Methods: The randomised clinical trial was conducted in Pondicherry Institute of Medical Sciences from September 2016 to March 2018, on 60 patients of either sex, aged between 20-60 years with American Society of Anesthesiologists (ASA) physical status I or II requiring elective intubations for general surgical procedures. The patients were randomly divided into two groups (n=30 each). Prior to induction group A received 1 µg/kg dexmedetomidine IV infusion over 10 min, and group B received 100 mL IV infusion of normal saline over 10 min. Also 2 minute before laryngoscopy group A received 10 mL of normal saline (IV) bolus, whereas, group B received esmolol 0.5 mg/kg IV diluted in 10 mL of normal saline as a bolus. Heart Rate (HR), Systolic Blood Pressure (SBP), Diastolic Blood Pressure (DBP) and Mean Arterial Pressure (MAP) were recorded at baseline i.e, pre-procedure before the study drug infusion, preinduction, at laryngoscopy (0min) and 1,3,5 min after intubation. A rise of MAP and/or HR more than 20% from the baseline was considered as positive laryngoscopy response. Student’s unpaired t-test was used for analysis of inter group variables. Intra group variables were analysed using repeated measures Analysis Of Variance (ANOVA). Results: Mean age of patients in group A was 38.77±13.082 years and group B was 37.20±13.069 years (p-value=0.644). Results revealed that both the groups had an increase in HR and MAP at 1 minute after laryngoscopy and intubation. Mean readings of MAP showed a maximum rise of group A (2.15%) vs group B (7.25%) from the baseline readings at 1min following laryngoscopy which showed no statistical significance. The maximum HR increase following laryngoscopy was at 1 minute in group A (8.28%) vs group B (13.59%), which were below the positive laryngoscopy response. The mean HR, SBP, DBP and MAP recorded at preinduction, at laryngoscopy, 1, 3 and 5 mins following intubation showed no statistical difference (p-value &gt;0.05) between the two groups. Conclusion: Usage of single dose preinduction dexmedetomidine iv infusion 1 µg/kg over 10 min was found to be equally effective in blunting the pressor response to laryngoscopy and intubation when compared to bolus dose of iv esmolol 0.5 mg/kg given 2 min prior to laryngoscopy.

Abelacimab Reduces Thrombus Propagation in a Baboon Model of Vascular Graft Thrombosis

Background: Factor XI (FXI) inhibition demonstrated strong efficacy in preventing thrombus formation in preclinical and clinical models of arterial and venous thrombosis. However, the effect of FXI inhibition in halting the progression of a formed clot remains largely unknown.Aims: This study aims to test whether abelacimab, a dual-acting FXI and activated FXI (FXIa) monoclonal antibody, is effective in halting clot formation and downstream growth when administered before or during active clot formation in an established baboon femoral arterio-venous (AV) shunt model.Methods: Three baboons had a chronic femoral AV shunt put in place; platelet and fibrin deposition inside and distal to collagen- or collagen + tissue factor (TF)-coated vascular grafts were measured at baseline (control), in a therapeutic setting, where abelacimab (1 mg/kg, intravenously) was administered 30 minutes after thrombus initiation, and in a preventative setting within the first 48 h and 1 week (144 - 216 h) post-administration. Pharmacodynamic effect was measured by activated partial thromboplastin time (aPTT).Results: Consistent with its half-life of 20 to 30 days, single iv administration of abelacimab at a dose of 1 mg/kg resulted in long-lasting (> 4-week) aPTT prolongation (> 2-fold). Administration of abelacimab 30 minutes after initiation of thrombosis using grafts coated either with collagen or with collagen + TF quickly halted downstream propagation of platelet and fibrin deposition compared to control. Further, downstream propagation of platelet and fibrin deposition was markedly reduced when clotting was induced by collagen, or collagen + tissue factor after abelacimab administration.Conclusions: These data suggest that abelacimab, a dual-acting anti-FXI/FXIa monoclonal antibody with a single long-lasting iv injection has the potential to slow down the growth and reduce the size of thrombi when admistered before or after clot induction. Data indicate a potential for therapeutic benefit of targeting FXI both in therapeutic and preventive settings.Sponsored by: Anthos Therapeutics Inc., 55 Cambridge Parkway, Suite 103, Cambridge, MA 02142 [Display omitted] DisclosuresWallisch: Aronora Inc,: Current Employment. Khder: Anthos Therapeutics: Consultancy; Novartis: Current equity holder in publicly-traded company, Other: Retiree. Bloomfield: Anthos Therapeutics: Current Employment. Gruber: Aronora Inc.: Current Employment, Current equity holder in publicly-traded company; Oregon Health and Science University: Current Employment.

A Novel Type V CRISPR System with Potential for Genome Editing in the Liver

RNA guided CRISPR genome editing systems can make specific changes to the genomes of mammalian cells and have the potential to treat a range of diseases including those that can be addressed by editing hepatocytes. Attempts to edit the liver in vivo have relied almost exclusively on the Cas9 nucleases derived from the bacteria S treptococcus pyogenes or Staphylococcus aureus to which humans are commonly exposed. Pre-existing immunity to both these proteins has been reported in humans which raises concerns about their in vivo application. In silico analysis of a large metagenomics database followed by testing in mammalian cells in culture identified MG29-1, a novel CRISPR system which is a member of the Type V family but exhibits only 41 % amino acid identity to Francisella tularensis Cas12a/cpf1. MG29-1 is a 1280 amino acid RNA programmable nuclease that utilizes a single guide RNA comprised of a 22 nucleotide (nt) constant region and a 20 to 25 nt spacer, recognizes the PAM KTTN (predicted frequency 1 in 16 bp) and generates staggered cuts. MG29-1 was derived from a sample taken from a hydrothermal vent and it is therefore unlikely that humans will have developed pre-existing immunity to this protein. A screen for sgRNA targeting serum albumin in the mouse liver cell line Hepa1-6 identified 6 guides that generated more than 80% INDELS. The MG29-1 system was optimized for in vivo delivery by screening chemical modifications to the guide that improve stability in mammalian cell lysates while retaining or improving editing activity. Two lead guide chemistries were evaluated in mice using MG29-1 mRNA and sgRNA packaged in lipid nanoparticles (LNP). Three days after a single IV administration on-target editing was evaluated in the liver by Sanger sequencing. The sgRNA that was the most stable in the in vitro assay generated INDELS that ranged from 20 to 25% while a sgRNA with lower in vitro stability failed to generate detectable INDELs. The short sgRNA and small protein size compared to spCas9 makes MG29-1 an attractive alternative to spCas9 for in vivo editing applications. Evaluation of the potential of MG29-1 to perform gene knockouts and gene additions via non-homologous end joining is ongoing. DisclosuresNo relevant conflicts of interest to declare.

Safe and Efficient Engraftment of CRISPR-Based ELANE Mono-Allelic Knocked out HSCs in Mice: Evidence for a Novel Treatment for ELANE Neutropenia

ELANE-related severe congenital neutropenia (SCN) is a disorder wherein numerous heterozygous mutations in the ELANE gene lead to misfolding and mislocalization of mutant neutrophil elastase, resulting in the death of myeloid cells and block in neutrophil differentiation.Currently, SCN is treated with daily injections of granulocyte colony-stimulating factor (G-CSF), but patients on this therapy are at risk of developing myelodysplastic syndrome (MDS) and acute myeloid leukemia (AML). The only other effective therapy is allogeneic hematopoietic stem cell transplantation that may involve graft versus host disease and risk of serious infections. Thus, there is great interest in safe and efficient therapeutic alternatives.Emendo Biotherapeutics has developed a novel CRISPR-based (OMNI A1) ex-vivo gene editing strategy that involves specific excision of disease-causing ELANE mutant allele. This treatment was tested on CD34 + HSCs from SCN patients and resulted in a significant improvement in the maturation of myeloid cells and their differentiation to normal functional neutrophils.To assess the feasibility and safety of this editing strategy, we conducted an in vivo study using human HSCs derived from two healthy donors that were engrafted into a NOD scid gamma (NSG) immunocompromised mice model. The study involved a single iv injection of HSCs that were either not-treated (NT), electroporated without delivery of CRISPR nuclease (mock) or edited with Emendo's novel nuclease, OMNI A1,(excision). Each group consisted of 6 NSG female mice and an additional group of 4 mice served as vehicle control (no HSCs). All animals were subjected to total body irradiation, 1 day prior to the treatment, at a dose of 250 cGy. Animals were subjected to retro-orbital blood collection, at 4 designated time points during the study (Figure A), for CD45 + cell counts using FACS. All animals were evaluated for systemic reactions, clinical signs and body weight changes during the study period. At the end of the study period, animals were subjected to peripheral blood (PB), bone marrow (BM) and spleen sampling. FACS analysis of CD45 + cells was performed on those samples to determine the engraftment efficiency. Furthermore, excision levels for ELANE were measured in the engrafted cells by droplet digital PCR (ddPCR). Finally, multilineage differentiation capacity of the engrafted cells was evaluated by FACS.Analysis of human CD45 + cells, at 20 weeks, showed comparable levels of engraftment of the not-treated, mock treated and excised HSCs in the mouse PB, BM and spleen (No statistically significant difference, Kruskal Wallis test) (Figure B). Excised cells were detected in the mouse BM and excision levels in some of the mice were comparable to those measured in the HSCs prior to engraftment (about 30-36%). Human-derived cells from the BM and spleen of mice engrafted with excised HSCs showed similar multilineage differentiation capacity as obtained in mice engrafted with either NT or mock-treated HSCs. Engrafted human CD45 + cells differentiated into neutrophils (CD66b +), myeloid cells (CD33 +), Lymphocytes: B-cells (CD19 +) and T-cells (CD3 +). In addition, a population of engrafted hematopoietic CD34 + cells was detected in the BM. Excised HSCs of both donors gave rise to all lineages tested, as efficient as the NT and the mock groups. Finally, engrafted mice showed no systemic reactions, abnormal clinical signs or loss of body weight.These data show that edited cells can be engrafted successfully, maintain their excision profile and populate the bone marrow. This study also supports the safety of the OMNI A1 therapeutic composition. [Display omitted] DisclosuresPovodovski: Emendo Biotherapeutic: Current Employment. Makaryan: Emendo Biotherapeutic: Research Funding. Sabo: Emendo Biotherapeutic: Research Funding. Dicken: Emendo Biotherapeutic: Ended employment in the past 24 months. Herman: Emendo Biotherapeutic: Current Employment. Emmanuel: Emendo Biotherapeutic: Current Employment. Dale: X4 Pharmaceuticals: Consultancy, Honoraria, Research Funding.

380 GS-3583, a novel FLT3 agonist Fc fusion protein, expands conventional dendritic cells in healthy volunteers

BackgroundConventional dendritic cells subtype 1 (cDC1) play a vital role in the priming and expansion of tumor specific CD8+ T cells and their recruitment to tumor microenvironment (TME). However, cDC1s are often underrepresented in the TME. Systemic administration of Fms-like tyrosine kinase 3 ligand (FLT3L), a hematopoietic growth factor that binds to FLT3 on myeloid and lymphoid progenitor cells, leads to expansion of cDC1s in the periphery which can then be recruited into the TME. FLT3 pathway stimulation using GS-3583, a novel FLT3 agonistic Fc fusion protein, has the potential to promote T cell mediated anti-tumor activity. We sought to evaluate the pharmacodynamic (PD) effect of a single dose of GS-3583 in healthy volunteers alongside its safety. Herein, we present the updated results of the study.MethodsThis was a first-in-human, placebo-controlled study of GS-3583 in healthy volunteers to evaluate the safety, pharmacokinetics (PK), and PD of escalating single doses (ranging from 75 micrograms to 2000 micrograms) of GS-3583. The study was blinded to the subjects and the investigator. Each dose cohort enrolled 8–12 healthy subjects who received GS-3583 or placebo as single IV infusion at 3:1 ratio. Subjects were observed in the phase 1 unit for 15 days and then for 12 weeks as outpatients. As part of the PD evaluation, we investigated the changes in the number of cDC1 and cDC2 cells.ResultsAs of 2nd July 2021, selected safety, PK and PD data from all 4 cohorts were available. GS-3583 was well tolerated and all subjects had been discharged. To date, there have been no serious or grade 3 or higher adverse events. Preliminary PK analysis suggested dose-dependent increase in GS-3583 exposure (AUC and Cmax). Preliminary PD analysis shows that administration of GS-3583 resulted in temporary, dose-dependent increases in cDC1/cDC2 cells that peaked between days 5–11 (higher doses resulted in later peaks) and returned to baseline within 3 weeks of drug administration (table 1, figure 1).Abstract 380 Table 1Selected subject characteristics and pharmacodynamic resultsAbstract 380 Figure 1A) Comparison of cDC1 cell quantitative changes in cohorts 1–4; B) Comparison of cDC2 cell quantitative changes in cohorts 1–4ConclusionsGS-3583 infusion was well tolerated and induced dose dependent expansion of dendritic cells in the periphery in healthy volunteers. In patients with cancer, this increase in dendritic cells can be utilized to enhance anti-tumor therapeutic effects of immuno-oncology therapies.AcknowledgementsFunding provided by Gilead Sciences, Inc.Ethics ApprovalThe study received study site IRB/Ethics Committee approval prior to enrollment of subjects.

The pharmacokinetics of letrozole and its effect on gonadotropins in anestrous ewes

The aim of this study was to determine the pharmacokinetics of letrozole and its effect on FSH and LH concentrations after single (IV, IM, SC) and repeated IV doses in anestrous ewes. This study was conducted in experiments 1 and 2 by randomly dividing 24 healthy Akkaraman ewes in anestrus into two equal groups. In experiment 1, the pharmacokinetics of letrozole following single IV, IM, and SC administration at 1 mg/kg dose and its effect of a single IV dose on plasma FSH and LH concentration were determined. In experiment 2, the effect of repeated IV doses of letrozole on FSH and LH concentrations was established. Plasma concentration of letrozole was measured using high-performance liquid chromatography, and pharmacokinetic parameters were calculated by non-compartmental analysis. FSH and LH concentrations were quantified using ELISA. The elimination half-life (t1/2ʎz) for IV, IM, and SC routes were 9.94, 37.29, and 41.07 h, respectively. The IV route for letrozole had a total clearance of 0.11 L/h/kg and a volume of distribution at a steady state of 1.50 L/kg. The peak plasma concentration was 0.11 μg/mL for the IM route and 0.14 μg/mL for the SC routes. The bioavailability was 55.18% for the IM route and 75.34% for the SC route. Letrozole following single and repeated (every 24 h for 3 days) IV administrations at 1 mg/kg dose did not affect LH concentration in anestrous ewes but caused an increase in the FSH concentration. This increase in FSH concentration may create a potential for the use of letrozole in ovarian superstimulation protocols. Favorable pharmacokinetic properties (long t1/2ʎz and good bioavailability) of letrozole for IM and SC routes require further investigation before use in estrus induction or estrus synchronization protocols in sheep.

Denosumab versus zoledronate for the treatment of low bone mineral density in male HIV-infected patients

IntroductionWe aimed to compare annual changes in the bone mineral density (BMD) at the lumbar spine (LS) and the femoral neck (FN) in males with HIV-associated osteoporosis treated with either zoledronate (ZOL) or denosumab (Dmab). MethodsIn this open label, 12-month, prospective, multicenter, cohort study, 23 male people living with HIV (PLWH) under antiretroviral therapy (ART) with low BMD were administered either a single iv infusion of ZOL 5 mg (n = 10) or Dmab 60 mg sc injections biannually (n = 13). Fourteen age-matched male PLWH with normal BMD served as controls. BMD was measured at baseline and at 12 months. ResultsLS-BMD increased within both treatment groups at 12 months (ZOL 5.43% ± 3.60%, p = 0.001; Dmab 5.76% ± 3.44%, p < 0.005) and decreased in controls (−2.58% ± 4.12, p = 0.04). FN-BMD increased in both treatment groups at 12 months (ZOL 7.23% ± 5.46%, p = 0.003; Dmab 3.01% ± 2.46%, p < 0.005), and remained unchanged in controls (1.22% ± 2.09, p = 0.06). LS-BMD changes did not differ between the two treatment groups, but FN-BMD changes were more prominent in the ZOL group (p < 0.05). None of our study cohort sustained new fragility fractures during the 12-month study period, and no case of acute phase response was recorded in the ZOL group. ConclusionsIn male PLWH under ART requiring osteoporosis treatment both ZOL and Dmab are efficient and well tolerated therapeutic options achieving BMD increases at least for the first year of treatment.

Abstract 917: The preclinical characterization of TST005, a bi-functional anti-PD-L1 and TGF-β trap fusion protein

Abstract Treatment with immune checkpoint inhibitors, including anti-PD(L)-1 antibodies, has demonstrated durable responses in patients with PD-L1 expressing tumors. However, not all patients with PD-L1 expression respond to the treatment due to de novo or acquired resistance to checkpoint inhibitors. The evidence from a number of studies showed that transforming growth factor β (TGF-β) signaling in the tumor microenvironment is associated with a poor response or resistance, and inhibition of TGF-β signaling can induce tumor T cell infiltration/activation and potentiate tumor response to immune checkpoint therapy. TST005 is a bifunctional fusion protein composed of the truncated extracellular domain of the TGF-βRII receptor (a TGF-β trap) fused to a humanized anti-PD-L1 IgG1 antibody (AM4B6 mAb) with ablated Fc immune effector function. TST005 bound to human/cyno PD-L1 with high affinity and blocked the PD-1/PD-L1 interaction potently in a cell-based reporter assay. Similarly, TST005 also bound to TGF-β1, 2 and 3 isoforms with high affinity and inhibited the activation of TGF-β receptor mediated signaling. In a superantigen stimulation assay with human peripheral blood mononuclear cells (PBMCs), TST005 enhanced T cell activation significantly as measured by interferon-γ (IFN-γ) production, as compared to TGF-β trap or AM4B6 mAb alone. In contrast, TST005 did not exert any cytokine release effect on human naïve PBMCs in vitro. TST005 showed a dose-proportional linear PK profile with single IV infusion in mice, and a negative correlation between plasma TST005 concentration and its pharmacodynamic marker TGF-β1 level was established. Furthermore, in the MC38/hPD-L1 xenograft mouse model, compared with TGF-β trap or AM4B6 mAb, TST005 increased the infiltration of activated CD8+ T cells into the tumor, and it resulted in tumor regression starting from 3mg/kg. Of particular interest, in the EMT6/hPD-L1 xenograft model which responds only moderately to PD-L1 inhibitors, TST005 induced significant tumor growth inhibition and regression starting from 10mg/kg. In conclusion, we have demonstrated that TST005 has enhanced immunomodulatory properties and can induce potent antitumor activity in preclinical tumor models that are not sensitive to PD-1/PD-L1 monotherapy. These results provide the rationale for further clinical evaluation of TST005 in patients with advanced solid tumors and less optimal response to first generation PD(L)-1 based immunotherapy. Citation Format: Hongjun Li, Chunming Wang, Huanhuan Guo, Xinlai Yao, Weiwei Sun, Di Sun, Yupeng Zhu, Xiangyu Xu, Zhenzhi Yan, Chengwei Jiang, Shengjie Zhang, Xiwen Wu, Yingdi Luo, Yadong Dai, Lisa Zheng, Fan Zhang, Fei Teng, Xueming Qian, Yi Gu. The preclinical characterization of TST005, a bi-functional anti-PD-L1 and TGF-β trap fusion protein [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 917.

Abstract 1747: Oncolytic Vesicular Stomatitis Virus expressing a species relevant interferon-beta is safe and provides a survival benefit when added as neoadjuvant to the standard of care for dogs with naturally occurring bone cancer

Abstract The goal of this study was to evaluate safety, efficacy, and immunological activity of Vesicular Stomatitis Virus (VSV) expressing a species relevant (canine) interferon-beta (IFN-β) and the sodium iodide symporter (NIS), henceforth called VSV, in companion dogs with naturally occurring appendicular osteosarcoma. The hypothesis was that VSV would track to the tumor, induce oncolysis, and lead to enhanced infiltration of immune cells, expansion of virus- and tumor specific T-cell clones, and improved survival outcomes.The first 15 dogs enrolled in the VSV Immunotherapy and Genomics of Osteosarcoma Research (VIGOR) study received a single iv dose of VSV (1x109 TCID50/kg) 10 days before amputation and adjuvant carboplatin chemotherapy. The final 13 dogs were randomized in a double-blinded fashion to receive either a single dose of VSV or placebo. Outcome data were compared to those from an historical control group of 57 dogs with appendicular osteosarcoma treated between 2006 and 2018 using the standard of care of alone. VSV treatment was associated with mild systemic effects (fever and inflammation), which were self-limiting and caused no symptomatology. Alterations in the tumor microenvironment (micronecrosis, increased fibrosis, and increased inflammation) were more common and more pronounced in VSV treated dogs than in placebo-treated dogs.Reduction in the expression of co-regulated cell cycle gene clusters in tumors after VSV-treatment was observed in dogs with longer survival. Conversely, increased expression of co-regulated immune/inflammatory gene clusters was observed in both VSV-treated dogs and in placebo-treated dogs, suggesting this might be, at least partly, due to the biopsy procedures. Assessment of changes in the clonal composition of T cells within the tumors, draining lymph nodes, and systemic circulation is in progress. No differences were evident in median event-free or overall survival times among the different groups, although aspects of the study design and sample size could obscure differences. Intriguingly, a “tail” of nine dogs (41%) that were long-term survivors (&amp;gt;450 days) was apparent in the VSV-treated group. All but one of these long-term survivors had evidence of inflammation in the pre-treatment tumor samples or showed increased inflammation in the post-treatment samples. We conclude that neoadjuvant VSV has an excellent safety profile and provides a survival benefit for a subset of dogs with spontaneous bone cancer. Ongoing work seeks to establish distinguishing characteristics that identify those individuals most likely to benefit from this therapy, as well as to evaluate the potential to enhance this benefit through the addition of novel immunomodulatory agents as part of our efforts to translate this approach to human cancer patients. Citation Format: Kelly M. Makielski, Aaron L. Sarver, Aishwarya Sathyanarayan, Michael S. Henson, Kathleen M. Stuebner, Alexandru-Flaviu Tabaran, Ingrid Cornax, Gerard O'Sullivan, Andrea Chehadeh, Donna Groschen, Kelly Bergsrud, Lauren J. Mills, Amber L. Winter, Sara Pracht, Milcah C. Scott, Michael A. Farrar, Gary R. Cutter, Joseph S. Koopmeiners, Stephen J. Russell, Shruthi Naik, Jaime F. Modiano. Oncolytic Vesicular Stomatitis Virus expressing a species relevant interferon-beta is safe and provides a survival benefit when added as neoadjuvant to the standard of care for dogs with naturally occurring bone cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1747.