Synthesis and in vitro biochemical evaluation of oxime bond-linked daunorubicin-GnRH-III conjugates developed for targeted drug delivery

Gonadotropin releasing hormone-III (GnRH-III), a native isoform of the human GnRH isolated from sea lamprey, specifically binds to GnRH receptors on cancer cells enabling its application as targeting moieties for anticancer drugs. Recently, we reported on the identification of a novel daunorubicin–GnRH-III conjugate (GnRH-III–[4Lys(Bu), 8Lys(Dau=Aoa)] with efficient in vitro and in vivo antitumor activity. To get a deeper insight into the mechanism of action of our lead compound, the cellular uptake was followed by confocal laser scanning microscopy. Hereby, the drug daunorubicin could be visualized in different subcellular compartments by following the localization of the drug in a time-dependent manner. Colocalization studies were carried out to prove the presence of the drug in lysosomes (early stage) and on its site of action (nuclei after 10 min). Additional flow cytometry studies demonstrated that the cellular uptake of the bioconjugate was inhibited in the presence of the competitive ligand triptorelin indicating a receptor-mediated pathway. For comparative purpose, six novel daunorubicin–GnRH-III bioconjugates have been synthesized and biochemically characterized in which 6Asp was replaced by D-Asp, D-Glu and D-Trp. In addition to the analysis of the in vitro cytostatic effect and cellular uptake, receptor binding studies with 125I-triptorelin as radiotracer and degradation of the GnRH-III conjugates in the presence of rat liver lysosomal homogenate have been performed. All derivatives showed high binding affinities to GnRH receptors and displayed in vitro cytostatic effects on HT-29 and MCF-7 cancer cells with IC50 values in a low micromolar range. Moreover, we found that the release of the active drug metabolite and the cellular uptake of the bioconjugates were strongly affected by the amino acid exchange which in turn had an impact on the antitumor activity of the bioconjugates

Decline in subarachnoid haemorrhage volumes associated with the first wave of the COVID-19 pandemic

BACKGROUND: During the COVID-19 pandemic, decreased volumes of stroke admissions and mechanical thrombectomy were reported. The study\u27s objective was to examine whether subarachnoid haemorrhage (SAH) hospitalisations and ruptured aneurysm coiling interventions demonstrated similar declines. METHODS: We conducted a cross-sectional, retrospective, observational study across 6 continents, 37 countries and 140 comprehensive stroke centres. Patients with the diagnosis of SAH, aneurysmal SAH, ruptured aneurysm coiling interventions and COVID-19 were identified by prospective aneurysm databases or by International Classification of Diseases, 10th Revision, codes. The 3-month cumulative volume, monthly volumes for SAH hospitalisations and ruptured aneurysm coiling procedures were compared for the period before (1 year and immediately before) and during the pandemic, defined as 1 March-31 May 2020. The prior 1-year control period (1 March-31 May 2019) was obtained to account for seasonal variation. FINDINGS: There was a significant decline in SAH hospitalisations, with 2044 admissions in the 3 months immediately before and 1585 admissions during the pandemic, representing a relative decline of 22.5% (95% CI -24.3% to -20.7%, p\u3c0.0001). Embolisation of ruptured aneurysms declined with 1170-1035 procedures, respectively, representing an 11.5% (95%CI -13.5% to -9.8%, p=0.002) relative drop. Subgroup analysis was noted for aneurysmal SAH hospitalisation decline from 834 to 626 hospitalisations, a 24.9% relative decline (95% CI -28.0% to -22.1%, p\u3c0.0001). A relative increase in ruptured aneurysm coiling was noted in low coiling volume hospitals of 41.1% (95% CI 32.3% to 50.6%, p=0.008) despite a decrease in SAH admissions in this tertile. INTERPRETATION: There was a relative decrease in the volume of SAH hospitalisations, aneurysmal SAH hospitalisations and ruptured aneurysm embolisations during the COVID-19 pandemic. These findings in SAH are consistent with a decrease in other emergencies, such as stroke and myocardial infarction

Development of mechanoreceptors.

The important morphogenetic and trophic interactions between axon terminals and their associated cells are widely studied. The first experiments examining this relationship were done by denervation studies in different types of mechanoreceptors. Not much was known about the precise nature of the factors, which may be involved in these interactions. The discovery of the nerve growth factor (NGF) made a breakthrough and the new expansion of this field of research started to focuse on bioactive agents and neurotrophic factors. The role of some of these new factors in the developing mechanoreceptors is described in this thesis. It focuses on the control factors of the cutaneous mechanoreceptors and proprioceptors development. It describes results of our studies of the Merkel cells developmental origin, the role of neurotrophins in their differentiation and maintenance, as well as our results concerning the role of ingrowing axons and the target tissue in the periphery on the number and structure of Herbst corpuscles and muscle spindles. In addition, the role of neurotrophins in Pacinian corpuscles and the role of transcription factors in the development of muscle spindles and Golgi tendon organs are described.Available from STL Prague, CZ / NTK - National Technical LibrarySIGLECZCzech Republi

A convenient synthetic method to improve immunogenicity of mycobacterium tuberculosis related T-cell epitope peptides

Epitopes from different proteins expressed by Mycobacterium tuberculosis (Rv1886c, Rv0341, Rv3873) were selected based on previously reported antigenic properties. Relatively short linear T-cell epitope peptides generally have unordered structure, limited immunogenicity, and low in vivo stability. Therefore, they rely on proper formulation and on the addition of adjuvants. Here we report a convenient synthetic route to induce a more potent immune response by the formation of a trivalent conjugate in spatial arrangement. Chemical and structural characterization of the vaccine conjugates was followed by the study of cellular uptake and localization. Immune response was assayed by the measurement of splenocyte proliferation and cytokine production, while vaccine e�cacy was studied in a murine model of tuberculosis. The conjugate showed higher tendency to fold and increased internalization rate into professional antigen presenting cells compared to free epitopes. Cellular uptake was further improved by the incorporation of a palmitoyl group to the conjugate and the resulted pal-A(P)I derivative possessed an internalization rate 10 times higher than the free epitope peptides. Vaccination of CB6F1 mice with free peptides resulted in low T-cell response. In contrast, significantly higher T-cell proliferation with prominent expression of IFN- , IL-2, and IL-10 cytokines was measured for the palmitoylated conjugate. Furthermore, the pal-A(P)I conjugate showed relevant vaccine e�cacy against Mycobacterium tuberculosis infection

Abstract 1122‐000243: Capacity and Characteristics of Thrombectomy Centers World‐wide Using the MT2020+ Global Thrombectomy Tracking Smartphone App

Introduction : Mechanical thrombectomy (MT) has been established as a first line therapy for large vessel occlusion stroke; however, MT remains underutilized globally with massive disparity in access based on country income level. Mission Thrombectomy 2020+ (MT2020+) is a global alliance and campaign that aims to reduce this disparity and democratize MT access for patients. A novel smartphone application, Global Thrombectomy Tracking App, was designed to characterize thrombectomy centers on a global‐level and numerically track MT cases in near real‐time. Methods : The MT2020 App was launched in November 2019. To gain insight into local systems of care, neurointerventionalists were prompted to participate in an optional 11‐question survey over a 19‐month period. Questions pertained to population density, organizational structure, academic affiliation, available imaging modalities, tPA usage, and case volumes. Results : Of 338 active users from 9 countries, 49‐neurointerventionalists participated in the survey. The majority (71.5%) practiced in large metropolis with population >1‐million, of which 16.3% were in mega‐cities (>10‐million). The centers were government funded (46.9%), private (40.8%) or charitable (12.2%). Most were academic hospitals (81.6%) with neurointerventional trainees (55.1%). At most centers (87.7%), IV‐tPA was available with annual treatment rate >5% for 55.1%. Most centers (57.1%) utilize additional CT perfusion scans prior to MT. For 69.3% respondents, the annual MT case volume was between 10–100 cases. Conclusions : Our survey analysis shows that the global MT tracking APP can generate important thrombectomy capacity and characteristics at regional levels on a global scale, which can be used for targeted funding and resource allocation to accelerate access to MT

Abstract Number ‐ 110: Case Report of Glue Embolism Complication after Lingual Artery Embolization and Rescue Thrombectomy

Introduction 68‐year‐old right‐handed female with right tongue‐base squamous cell carcinoma status‐post wide surgical‐excision and flap reconstruction, whopresented with large volume right‐sided orolingual hemorrhage. Due to concerns of impending airway compromise and a history of difficult intubation, Interventional Neuroradiology was consulted to perform an angiogram and embolization. Methods A 6FrEnvoy was advanced into the external carotid artery and the origin of the lingual artery. Despite repeated attempts using a variety of microsystems, distal access within the lingual artery could not be achieved (Fig. 1B). N‐butyl‐2‐cyanoacrylate was prepared and injected with adequate penetration. Negative pressure was applied to the microcatheter and withdrawn. The guide catheter was aspirated and rapidly withdrawn under negative pressure. A control angiogram was being prepared when an intracranial glue cast was seen and confirmed to be a proximal M2‐occlusion. 4000U of Heparin werebolusedat this time. An Advantage Exchange wire facilitated an exchange for an 8Fr groin sheath and a Walrus catheter. The glue was crossed with a microsystem microsystem and a Solitaire‐4×40mm was deployed. A ZOOM55 was advanced to the face of the glue caste and together these were pulled into the Walrus catheter. Serial angiograms revealed stenosis, but patency. Results Initial post‐thrombectomy exam revealed mild left‐hand paresis. However, the patient’s neurologic status declined and progressed to left arm hemiparesis, facial droop, and right gaze‐preference. An angiogram was performed, which revealed re‐occlusion of the left M2 trunk. Using aTrevo‐3×32mmstentriever, the M2‐occlusion was recanalized with persistent stenosis. Serial angiograms were concerning for progressing re‐occlusion. The patient improved with residual distal left hemiparesis. Conclusions •Distal access may be extremely difficult in select cases. The proximal location of the microcatheter relative to the origin of the lingual artery and the external carotid artery off the common carotid artery, likely contributed to the embolic event. The use of progressively smaller microcatheters to facilitate distal access in turn biased the choice of embolization materials. Lastly, there was evidence of vasospasm in the lingual artery, which may have contributed to proximal polymerization and adherence to the catheters during their removal. •After successful retrieval of the glue material, there was residual stenosis of unclear etiology, possibly due to denuded endothelium, dissection, or residual glue promoting in‐situ thrombosis. The eventual clearing of thebolusedheparin may have contributed to the delayed re‐occlusion. Recanalization after re‐occlusion was only possible after thrombectomy. •The use of an intermediate catheter may mitigate the risk of glue embolization by facilitating distal access and/or by providing an additional conduit for aspirating refluxed glue. If distal access into a culprit vessel can not be achieved, coil embolization may be a safer choice. Re‐canalized glue embolisms may be prone to re‐occlusion. Continuing a heparin drip may be prudent practice to prevent re‐occlusion

Critical Angiographic and Sonographic Analysis of Intra Aneurysmal and Downstream Hemodynamic Changes After Flow Diversion

IntroductionSuccessful treatment of intracranial aneurysms after flow diversion (FD) is dependent on the flow modulating effect of the device. We aimed to investigate the intra-aneurysmal and parent vessel hemodynamic changes, as well as the incidence of silent emboli following treatment with various FD devices.MethodsWe evaluated the appearance of the eclipse sign in nine distinct phases of cerebral angiography before and immediately after FD placement in correlation with aneurysm occlusion. Angiographic and clinical data of consecutive procedures were analyzed retrospectively. Patients who had successful FD procedure without adjunctive coiling, visible eclipse sign on post embolization angiography, and reliable follow-up angiographic data were included in the analysis. Detailed analysis of hemodynamic data from transcranial doppler after FD was performed in selected patients, such as monitoring for silent emboli.ResultsAmong all patients (N = 65) who met inclusion criteria, complete aneurysm occlusion at 12 months was achieved in 89% (58/65). Eclipse sign prior to FD was observed in 42% (27/65) with unchanged appearance in 4.6% (3/65) of the treated patients. None of these three patients achieved complete aneurysm occlusion. Among all analyzed variables, such as aneurysm size, device type used, age, and appearance of the eclipse sign pre- and post-FD, the most reliable predictor of permanent aneurysm occlusion at 12 months was earlier, prolonged, and sustained eclipse sign visibility in more than three angiographic phases in comparison to the baseline (p < 0.001). Elevation in flow velocities within the ipsilateral vascular territory was noted in 70% (9/13), and bilaterally in 54% (7/13) of the treated patients. None of the patients had silent emboli.ConclusionsIntra-aneurysmal and parent vessel hemodynamic changes after FD can be reliably assessed by the cerebral angiography and transcranial doppler with important implications for the prediction of successful treatment

Abstract 1122‐000121: Integrated Geomapping Tool of Certified Stroke Centers in United States: A SVIN MT2020+ Committee Collaboration

Introduction: Stroke is a leading cause of morbidity, mortality and healthcare spending in the United States. Acute management of ischemic stroke is time‐dependent and evidence suggests improved clinical outcomes for patients treated at designated certified stroke centers. There is an increasing trend among hospitals to obtain certification as designated stroke centers. A common source or integrated tool providing both information and location of all available stroke centers in the US irrespective of the certifying organization is not readily available. The objective of our research is to generate a comprehensive and interactive electronic resource with combined data on all geographically‐coded certified stroke centers to assist in pre‐hospital triage and study healthcare disparities in stroke including availability and access to acute stroke care by location and population. Methods: Data on stroke center certification was primarily obtained from each of the three main certifying organizations: The Joint Commission (TJC), Det Norske Veritas (DNV) and Healthcare Facilities Accreditation Program (HFAP). Geographic mapping of all stroke center locations was performed using the ArcGIS Pro application. The most current data on stroke centers is presented in an interactive electronic format and the information is frequently updated to represent newly certified centers. Utility of the tool and its analytics are shown. Role of the tool in improving pre‐hospital triage in the stroke systems of care, studying healthcare disparities and implications for public health policy are discussed. Results: Aggregate data analysis at the time of submission revealed 1,806 total certified stroke centers. TJC‐certified stroke centers represent the majority with 106 Acute Stroke Ready (ASR), 1,040 Primary Stroke Centers (PSCs), 49 Thrombectomy Capable Centers (TSCs) and 197 Comprehensive Stroke Centers (CSCs). A total of 341 DNV‐certified programs including 36 ASRs, 162 PSCs, 16 PSC Plus (thrombectomy capable) and 127 CSCs were identified. HFAP‐certified centers (75) include 16 ASRs, 49 PSCs, 2 TSCs and 8 CSCs. A preliminary map of all TJC‐certified CSCs and TSCs is shown in the figure (1). Geospatial analysis reveals distinct areas with currently limited access to certified stroke centers and currently, access to certified stroke centers is extremely limited to non‐existent in fe States (for example: Idaho, Montana, Wyoming, New Mexico and South Dakota). Conclusions: Stroke treatment and clinical outcomes are time‐dependent and prompt assessment and triage by EMS directly to appropriate designated stroke centers is therefore critical. A readily available electronic platform providing location and treatment capability for all nearby certified centers will enhance regional stroke systems of care, including enabling more rapid inter‐hospital transfers for advanced intervention. Identifying geographic areas of limited access to treatment can also help improve policy and prioritize the creation of a more equitable and well‐distributed network of stroke care in the United States

Aneurysmal subarachnoid hemorrhage in pregnancy: National trends of treatment, predictors, and outcomes.

IntroductionAneurysmal subarachnoid hemorrhage (aSAH) is a rare event associated with significant pregnancy-associated maternal and neonatal morbidity and mortality. The optimal treatment strategy and clinical outcome of aSAH in pregnancy remains unclear. We aimed to investigate the treatment utilizations and outcomes of aSAH in pregnant people.MethodsUsing the 2010-2018 National Inpatient Sample, we identified all birth hospitalizations of women between ages of 18 to 45 associated with subarachnoid hemorrhage and aneurysm treatment were included. Multivariate analyses were used to evaluate the effect of pregnancy state, mode of treatment of aneurysms, severity of subarachnoid hemorrhage on mortality and discharge destination of this cohort. Trends in mode of treatment utilized for aneurysmal treatment in this time interval was evaluated.Results13,351 aSAH with treatment were identified, of which 440 were associated with pregnancy. There was no significant difference in mortality or rate of discharge to home in pregnancy related hospitalization. Worse aSAH severity, chronic hypertension, and smaller hospital size was associated with significantly higher rate of mortality from aSAH during pregnancy. Worse aSAH severity was associated with lower rate of discharge to home. Like the non-pregnant cohort, the treatment of ruptured aneurysms in pregnancy are increasingly through endovascular approaches. The mode of treatment does not change the mortality or discharge destination.ConclusionsPregnancy does not alter mortality or the discharge destination for aSAH. Ruptured aneurysms during pregnancy are increasingly treated endovascularly. Mode of aneurysm treatment does not affect mortality or discharge destination in pregnancy