Impact of SARS-CoV-2 Pandemic on "Stroke Code" Imaging Utilization and Yield.

BACKGROUND AND PURPOSE: Indirect consequences of the Severe Acute Respiratory Syndrome coronavirus 2 (SARS-CoV-2) pandemic include those related to failure of patients to seek or receive timely medical attention for seemingly unrelated disease. We report our experience with stroke code imaging during the early pandemic months of 2020. MATERIALS AND METHODS: Retrospective review of stroke codes during the 2020 pandemic and both 2020 and matched 2019 prepandemic months was performed. Patient variables were age, sex, hospital location, and severity of symptoms based on the NIHSS. We reviewed the results of CT of the head, CTA, CTP, and MR imaging examinations and classified a case as imaging-positive if any of the imaging studies yielded a result that related to the clinical indication for the study. Both year-to-year and sequential comparisons were performed between pandemic and prepandemic months. RESULTS: A statistically significant decrease was observed in monthly stroke code volumes accompanied by a statistically significant increased proportion of positive imaging findings during the pandemic compared with the same months in the prior year (P < .001) and prepandemic months in the same year (P < .001). We also observed statistically significant increases in average NIHSS scores (P = .045 and P = .03) and the proportion of inpatient stroke codes (P = .003 and P = .03). CONCLUSIONS: During our pandemic period, there was a significantly decreased number of stroke codes but simultaneous increases in positivity rates, symptom severity, and inpatient codes. We postulate that this finding reflects the documented reluctance of patients to seek medical care during the pandemic, with the shift toward a greater proportion of inpatient stroke codes potentially reflecting the neurologic complications of the virus itself.

Multidimensional characterization of carotid artery stenosis using CT imaging: a comparison with ultrasound grading and peak flow measurement.

PURPOSE: Clinical decision making for carotid surgery depends largely upon stenosis grade. While digital subtraction angiography remains the gold standard for stenosis grading, many physicians use less invasive modalities. The purpose of this study was to compare the results of multidimensional Computed tomography (CTA) with ultrasound (US) grading and peak flow velocity (PSV). METHODS: 37 stenosed carotid arteries were studied retrospectively in 36 consecutive patients. US grading and PSV were compared to multidimensional CTA analysis (diameter, area and volumetric measurements), performed by a medical software company. Calculations of stenosis percentage on CTA were made using the NASCET and ECST methodology. Diameter measurements were also performed by a neuroradiologist. RESULTS: All CTA diameter, area and volume measurements had only modest correlation with PSV (r<0.5) and ultrasound grading (p<0.5). There was concordant classification of stenosis grades in only 40-60% of cases. CTA diameter, area and volume measurements had good correlation (0.69

Accuracy of the Alberta Stroke Program Early CT Score during the first 3 hours of middle cerebral artery stroke: comparison of noncontrast CT, CT angiography source images, and CT perfusion.

BACKGROUND AND PURPOSE: The Alberta Stroke Program Early CT Score (ASPECTS) is a reliable method of delineating the extent of middle cerebral artery (MCA) stroke. Our aim was to retrospectively compare the accuracy of ASPECTS on noncontrast CT, CT angiography (CTA) source images, and CT perfusion maps of cerebral blood volume (CBV) during the first 3 hours of middle cerebral artery (MCA) stroke. MATERIALS AND METHODS: First-time patients with MCA stroke who presented <3 hours from symptom onset and were evaluated by noncontrast CT/CTA/CT perfusion, had confirmed acute nonlacunar MCA infarct on diffusion-weighted MR imaging (DWI) within 7 days, and had follow-up angiography were included. Patients were excluded for persistent MCA occlusion or stenosis. Two raters through consensus assigned an ASPECTS on the noncontrast CT, CTA source images, and the section-selective (2 x 12 mm coverage) CT perfusion CBV maps. ASPECTS on follow-up DWI served as the reference standard. For each CT technique, the detection rates of regional infarction, the mean ASPECTS, and the linear correlation to final ASPECTS were determined and compared. P values <.05 were considered significant. RESULTS: Twenty-eight patients satisfied the criteria with DWI performed at a mean of 50.3 hours (range, 22-125 hours) post-CT imaging. Of 280 ASPECTS regions, 100 were infarcted on DWI. The accuracy of noncontrast CT, CTA source images, and CT perfusion CBV for detecting regional infarct was 80.0%, 84.3%, and 96.8%, respectively (P < .0001). The mean ASPECTSs of noncontrast CT, CTA source images, CT perfusion CBV, and DWI were 8.4 +/- 1.8, 8.0 +/- 1.8, 6.8 +/- 1.9, and 6.5 +/- 1.8, respectively. The mean noncontrast CT and CTA source image ASPECTS was different from that of DWI (P < .05). Correlation of noncontrast CT, CTA source images, and CT perfusion CBV ASPECTS with final ASPECTS was r(2) = 0.34, r(2) = 0.42, and r(2) = 0.91, respectively. CONCLUSION: In a retrospective cohort of MCA infarcts imaged <3 hours from stroke onset, ASPECTS was most accurately determined on CT perfusion CBV maps.

Measuring elevated microvascular permeability and predicting hemorrhagic transformation in acute ischemic stroke using first-pass dynamic perfusion CT imaging.

BACKGROUND AND PURPOSE: Hemorrhagic transformation (HT) can be a devastating complication of acute ischemic stroke (AIS). The purpose of this study was to determine whether increased microvascular permeability (PS) of the blood-brain barrier was detected in early AIS by using first-pass dynamic perfusion CT (PCT) and whether PS was significantly higher in infarcts destined for HT. MATERIALS AND METHODS: Fifty patients with AIS less than 3 hours old and evaluated by PCT were included. PS color maps were retrospectively generated from PCT data using the Patlak model. One reader analyzed each PS map by drawing 4 circular 10-mm regions of interest on any focal abnormality. The mean of these 4 regions of interest represented the PS of the infarct (PSinfarct). The mean of 4 mirror regions of interest on the nonischemic contralateral hemisphere was also obtained (PScontrol). PSinfarct and PScontrol were compared by using an exact Wilcoxon test. PSinfarct for infarcts that developed HT on follow-up (PSHT) was compared with all of the others (PSNo-HT) using an exact Mann-Whitney test. RESULTS: Forty-four infarcts (88%) showed focal PS elevation in the region of infarct. In units of milliliters per 100 milliliters per minute, PSinfarct ranged from 0 to 13 (mean: 3.5+/-3.1) versus PScontrol of 0-0.8 (mean: 0.28+/-0.27; P<.0001). Six infarcts (12%) developed HT, all of which were within the region of PS elevation. PSHT ranged from 5.2 to 13 (mean: 9.8+/-2.9) versus PSNo-HT of 0-5.9 (mean: 2.7+/-2.0; P<.0001). Eighteen infarcts (36%) were treated with recombinant tissue plasminogen activator (rtPA). A significant difference between PSHT and PSNo-HT persisted irrespective of rtPA treatment. CONCLUSIONS: Elevated permeability was detectable in AIS by using first-pass PCT and it predicted subsequent HT.

Dynamic sagittal half-Fourier acquired single-shot turbo spin-echo MR imaging of the temporomandibular joint: initial experience and comparison with sagittal oblique proton-attenuation images.

BACKGROUND AND PURPOSE: Our aim was to assess dynamic half-Fourier acquired single-shot turbo spin-echo (HASTE) MR imaging of the temporomandibular joint (TMJ) using parallel imaging, in comparison with static proton density (Pd) imaging. MATERIALS AND METHODS: Thirty-four TMJs from 17 subjects (7 volunteers, 10 patients) were imaged in a multichannel head coil on a 1.5 T magnet by using a 35-second dynamic sagittal HASTE acquisition (TR/TE, 1180/65 msec; matrix, 128 x 128; section thickness, 7 mm; 30 images) and sagittal oblique Pd in closed- and open-mouthed positions (TR/TE, 1800/12 msec; matrix, 256 x 256; section thickness, 2 mm; 15 sections). Images were reviewed by 3 readers and rated for confidence of disk position, presence of motion artifact, range of motion, and presence of disk displacement on a 5-point scale. Consensus review of cases was also performed to assess disk dislocation and limited range of motion. RESULTS: More static examinations were rated as having motion artifact (19.6% versus 6.9%, P=.016), limited range of motion (30.4% versus 17.7%, P=.016), and disk dislocations (31.4% versus 22.6%, P=.071). Confidence ratings were higher on dynamic examinations (4.11 versus 3.74, P=.018). Chi-squared tests demonstrated no significant difference in consensus reviews of the 2 examination types. CONCLUSION: Dynamic HASTE TMJ MR imaging is a time-efficient adjunct to standard MR imaging protocols, producing fewer motion artifacts, additional range of motion information, and a dynamic assessment of disk position, when compared with static imaging. Further study is needed to evaluate the role of this sequence in diagnosing disk displacement.

Sch 486058: a novel cyclic peptide of actinomycete origin.

The structure of a novel cyclic peptide (1) produced by Actinomycete sp. has been assigned on the basis of extensive NMR and mass spectral data.

Carboxyalkylated histidine is a pH-dependent product of pegylation with SC-PEG.

PURPOSE: Pegylation of therapeutic protein usually results in a mixture of monopegylated proteins with differing sites of modification. With rh-interferon-alpha2A pegylation, we have found that this heterogeneity includes two classes of pegylation site chemistry, the relative proportions of which can be adjusted by reaction pH. METHODS: The effect of pegylation reaction pH on the relative proportion of three peaks produced was investigated. Products were purified and characterized by peptide mapping, chemical stability to neutral hydroxylamine, and biologic activity. RESULTS: Reactions at basic pH levels produced a mixture of products pegylated at lysine residues as has been observed elsewhere. However, the dominant product of reactions at mildly acidic levels of pH showed distinct chemistry and higher cytopathic effect activity. The primary site of modification at this pH was His34. We developed a quantitative assay using sensitivity to neutral hydroxylamine to measure the proportion of urethane bonds involving carboxyalkylated histidines. This assay showed that histidine was pegylated preferentially at low pH levels with another protein, rh-Interleukin-10. CONCLUSIONS: Reaction pH can be used to select the preferred pegylation site chemistry.

Fellowship and practice trends in neuroradiology training programs in the United States.

BACKGROUND AND PURPOSE: Neuroradiology has become an increasingly diverse and subspecialized discipline. We evaluated the current status and trends affecting fellowship programs and the practice of clinical neuroradiology at academic medical centers, with emphasis on invasive procedures. METHODS: All 85 program directors at Accreditation Council for Graduate Medical Education-approved fellowships in neuroradiology were sent a detailed questionnaire pertaining to various demographic aspects of their program and the performance of certain radiologic examinations of the brain and spine. RESULTS: Sixty-seven programs (79%) responded. As many as 50% of programs are 1 year in length. Twenty-five percent of 2-year fellows leave their program after 1 year of training. During the past 5 years, 36% of programs have decreased in size and 73% reported a decline in the number of applicants. The majority (55%) of programs have had applicants renege on their commitment to begin a fellowship. Twenty percent of 2-year programs do not offer training in endovascular interventional procedures. Neurosurgeons perform endovascular interventional procedures at 18% of centers. There is an 18-fold variation in the volume of neuroangiographic procedures performed each year and a 150-fold variation in the volume of myelographic procedures performed. In 29% of programs, neuroradiologists are nonparticipants in nonvascular interventional spinal procedures; in 40%, they share these procedures with musculoskeletal radiologists/nonradiologists. CONCLUSION: Interest in fellowship programs in neuroradiology is declining. An applicant's commitment to either begin a fellowship or complete 2 years of training cannot be regarded with assurance, and there is a lack of uniformity in many areas of the training experience, particularly in invasive diagnostic and therapeutic procedures.

Structural characterization of the oxidative degradation products of an antifungal agent SCH 56592 by LC-NMR and LC-MS.

LC-NMR and LC-MS were used to characterize the structures of four major degradation products of SCH 56592, an antifungal drug candidate in clinical trials. These compounds were formed under stress conditions in which the bulk drug substance was heated in air at 150 degrees C for 12 days, and were separated from SCH 56592 as a mixture using a semi-preparative HPLC method. The data from LC-NMR, LC-ESI-MS (electrospray ionization mass spectrometry) and LC-ESI-MS/MS indicate that the oxidation occurred at the piperazine ring in the center of the drug molecule. The structures of the degradation products were determined from the 1H NMR spectra obtained via LC-NMR, which were supported by LC-ESI-MS and LC-ESI-MS/MS analyses. A novel degradation pathway of SCH 56592 was proposed based on these characterized structures.

Biochemical characterization of the gamma-secretase activity that produces beta-amyloid peptides.

Recent studies of gamma-secretase have pointed out that it may be comprised of a multisubunit complex with presenilin 1 and presenilin 2 as central components. Elucidation of the biochemical mechanism of this enzymatic activity will provide important information for developing gamma-secretase inhibitors in Alzheimer's disease therapy. Here we describe the biochemical characterization of gamma-secretase activities using a sensitive, membrane-based assay system. Membranes were isolated from 293 cells expressing C99, the substrate of gamma-secretase. Upon incubation at 37 degrees C, C99 is cleaved by the endogenous gamma-secretase, and Abeta peptides are liberated. Abeta40 and Abeta42 gamma-secretase activities are very similar in terms of their kinetic profiles and pH dependence, supporting the notion that a single enzyme is involved in both Abeta40 and Abeta42 production. Pepstatin A inhibited Abeta40 and Abeta42 gamma-secretase activities with similar potency. Peptide difluoroketone and peptide aldehyde inhibitors inhibited Abeta40 production in a dose-dependent fashion, enhanced Abeta42 production at low concentrations, and inhibited Abeta42 production at high concentrations. Although the selective increase of Abeta42 by low concentrations of peptide difluoroketone and peptide aldehyde inhibitors has been reported in intact cells, the finding that this phenomenon occurs in a membrane-based assay system suggests that these compounds increase Abeta42 by a direct effect on gamma-secretase. The ability of these compounds to increase Abeta42 production may reflect allosteric modulation of the gamma-secretase complex by a mechanism related to that responsible for the increase of Abeta42 production by mutations in presenilins.

ADP is the cognate ligand for the orphan G protein-coupled receptor SP1999.

P2Y receptors are a class of G protein-coupled receptors activated primarily by ATP, UTP, and UDP. Five mammalian P2Y receptors have been cloned so far including P2Y1, P2Y2, P2Y4, P2Y6, and P2Y11. P2Y1, P2Y2, and P2Y6 couple to the activation of phospholipase C, whereas P2Y4 and P2Y11 couple to the activation of both phospholipase C and the adenylyl cyclase pathways. Additional ADP receptors linked to Galpha(i) have been described but have not yet been cloned. SP1999 is an orphan G protein-coupled receptor, which is highly expressed in brain, spinal cord, and blood platelets. In the present study, we demonstrate that SP1999 is a Galpha(i)-coupled receptor that is potently activated by ADP. In an effort to identify ligands for SP1999, fractionated rat spinal cord extracts were assayed for Ca(2+) mobilization activity against Chinese hamster ovary cells transiently transfected with SP1999 and chimeric Galpha subunits (Galpha(q/i)). A substance that selectively activated SP1999-transfected cells was identified and purified through a series of chromatographic steps. Mass spectral analysis of the purified material definitively identified it as ADP. ADP was subsequently shown to inhibit forskolin-stimulated adenylyl cyclase activity through selective activation of SP1999 with an EC(50) of 60 nM. Other nucleotides were able to activate SP1999 with a rank order of potency 2-MeS-ATP = 2-MeS-ADP > ADP = adenosine 5'-O-2-(thio)diphosphate > 2-Cl-ATP > adenosine 5'-O-(thiotriphosphate). Thus, SP1999 is a novel, Galpha(i)-linked receptor for ADP.

High-resolution LC/MS for analysis of minor components in complex mixtures: negative ion ESI for identification of impurities and degradation products of a novel oligosaccharide antibiotic.

High-resolution mass spectrometry has been routinely used for structural confirmation and identification; however, it has mostly been applied to relatively pure samples. Exact mass measurement of minor components such as impurities, degradation products or metabolites in complex mixtures has been difficult without prior separation and isolation. Here we report the utilization of on-line liquid chromatography in combination with high-resolution mass spectrometry for the identification of impurities and base degradation products of Sch 27899, a member of the everninomicin class of antibiotics. Nine Sch 27899-related impurities and degradation products were detected by negative ion electrospray ionization using a magnetic sector mass spectrometer. Exact mass measurements were obtained at a resolution of 5000 using polyethylene glycol (PEG) sulfates as internal standards. Corresponding elemental compositions were determined within a 2 ppm error tolerance and structures were proposed for all components.

Extraction and characterization of adenovirus proteins from sodium dodecylsulfate polyacrylamide gel electrophoresis by matrix-assisted laser desorption/ionization mass spectrometry.

A new methodology for the extraction and characterization of proteins from Coomassie-stained sodium dodecylsulfate polyacrylamide gel electrophoresis using matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) has been described. The utility of this methodology was demonstrated in the characterization of adenovirus proteins. The key steps in the extraction and destaining process involve washing the excised band with a combination of solvents that include 10% acetic acid, acetonitrile, methanol, and formic acid:water:isopropanol mixture. By using this procedure, we determined adenovirus proteins with molecular weights ranging from 10,000 to 110,000 Da by MALDI-MS, obtaining a detection limit of approximately 6 pmol. Parallel experiments were successfully carried out to analyze adenovirus proteins from Cu-stained gels. It was observed that increase in laser intensity resulted in significant improvements in the quality of MALDI mass spectra for the analysis of inefficiently destained proteins from Cu-stained gels.

Mass spectrometric mapping of disulfide bonds in recombinant human interleukin-13.

Interleukin 13 (IL-13), a member of the a-helical family of cytokines, has approximately 30% primary sequence homology with IL-4 and shares a common receptor component. The biologically active rhIL-13 is monomeric and non-glycosylated, and contains two disulfide bonds as determined by comparative electrospray mass spectrometric (MS) analysis of the protein before and after reduction with dithiothreitol-dithioerythritol. A trypsin-resistant core peptide of rhIL-13 was isolated and analyzed by plasma desorption (PD) MS, identifying a disulfide-linked core peptide. Subsequent digestion of this core peptide by pepsin, followed by PDMS analysis of the resulting cystine-containing peptic fragments, provided rapid determination of the existing disulfide bonds between cysteine residues 28-56 and 44-70. This disulfide arrangement is similar to that observed for the analogous four internal cysteine residues in hIL-4. The conservation of disulfide bond arrangements between hIL-13 and hIL-4, coupled with their alpha-helical structure and sequence homologies, confirms that IL-13 and IL-4 are structural homologues. It is also consistent with their reported similarities in biological function and receptor binding kinetics.

Identification of pharmacokinetically stable 3, 10-dibromo-8-chlorobenzocycloheptapyridine farnesyl protein transferase inhibitors with potent enzyme and cellular activities.

Farnesyl protein transferase (FPT) is a promising target for the development of cancer chemotherapeutics because it is responsible for the farnesylation of oncogenic p21 Ras proteins which are found in nearly 30% of all human cancers and necessary for cellular development and growth. The recent discovery and progression to phase II clinical trials of trihalobenzocycloheptapyridine Sch-66336 as a potent inhibitor of FPT with oral, in vivo efficacy in mice have spawned extensive structure-activity relationship studies (SAR) of this class of compounds. Of the many trihalobenzocycloheptapyridine analogues prepared, we have identified several which inhibit FPT and cellular proliferation at single-digit nanomolar concentrations and which have good pharmacokinetic properties in mice.

Waxing and waning gynecomastia: an indication of noncompliant use of prescribed medication.

We present two cases of recurrent gynecomastia in men enrolled in a placebo-controlled trial evaluating the efficacy of finasteride in treating benign prostatic hyperplasia. When the pharmacologic records were examined, it was apparent that the breast tissue hyperplasia diminished when the patients become noncompliant with their study medication and then resumed therapy. Because of the difficulty in obtaining accurate data on an individual's ability to maintain a consistent pharmacologic regimen, we believe that observing such "waxing and waning gynecomastia" may provide the physician with a clue regarding a patient's actual compliance with certain medications.

Isolation and characterization of novel human immunodeficiency virus integrase inhibitors from fungal metabolites.

We have identified a series of novel inhibitors of human immunodeficiency virus type 1 (HIV-1) integrase by randomly screening natural product extracts using an in vitro biochemical assay designed to identify inhibitors of integrase-catalysed strand transfer. Equisetin recovered from the fungus Fusarium heterosporum and a novel enantiomeric homologue of equisetin from Phoma sp. were isolated as inhibitors of HIV-1 integrase in vitro. Two additional analogues, a novel decalin derivative, integric acid, and oteromycin were also discovered to be inhibitors of integrase. Equisetin and related compounds inhibit 3' end-processing and strand transfer as well as disintegration catalysed by either the full-length enzyme or the truncated integrase core domain (amino acids 50-212). These compounds also inhibit strand transfer reactions catalysed by stable complexes assembled in vitro and integration reactions catalysed by pre-integration complexes isolated from HIV-1-infected cells. The compounds described in this report are structurally novel and mechanistically distinct from many previously described inhibitors of HIV-1 integrase. These results demonstrate the utility of using an appropriately configured assay to identify compounds that are effective post-assembly and the potential of isolating novel integrase inhibitors from complex natural product extracts.

A glucan synthase FKS1 homolog in cryptococcus neoformans is single copy and encodes an essential function.

Cryptococcal meningitis is a fungal infection, caused by Cryptococcus neoformans, which is prevalent in immunocompromised patient populations. Treatment failures of this disease are emerging in the clinic, usually associated with long-term treatment with existing antifungal agents. The fungal cell wall is an attractive target for drug therapy because the syntheses of cell wall glucan and chitin are processes that are absent in mammalian cells. Echinocandins comprise a class of lipopeptide compounds known to inhibit 1,3-beta-glucan synthesis, and at least two compounds belonging to this class are currently in clinical trials as therapy for life-threatening fungal infections. Studies of Saccharomyces cerevisiae and Candida albicans mutants identify the membrane-spanning subunit of glucan synthase, encoded by the FKS genes, as the molecular target of echinocandins. In vitro, the echinocandins show potent antifungal activity against Candida and Aspergillus species but are much less potent against C. neoformans. In order to examine why C. neoformans cells are less susceptible to echinocandin treatment, we have cloned a homolog of S. cerevisiae FKS1 from C. neoformans. We have developed a generalized method to evaluate the essentiality of genes in Cryptococcus and applied it to the FKS1 gene. The method relies on homologous integrative transformation with a plasmid that can integrate in two orientations, only one of which will disrupt the target gene function. The results of this analysis suggest that the C. neoformans FKS1 gene is essential for viability. The C. neoformans FKS1 sequence is closely related to the FKS1 sequences from other fungal species and appears to be single copy in C. neoformans. Furthermore, amino acid residues known to be critical for echinocandin susceptibility in Saccharomyces are conserved in the C. neoformans FKS1 sequence.

The role of mass spectrometry in the drug discovery process.

The structure characterization of biologically-active organic compounds, developed from synthetic and natural sources, is an integral part of the drug discovery effort to identify novel therapeutic agents. Mass spectrometric methods (electrospray ionization, matrix-assisted laser desorption/ionization, fast atom bombardment, electron ionization and chemical ionization) are uniquely qualified to solve a wide variety of structural identification problems with high speed and accuracy. This report provides an overview of the recent developments in mass spectrometry (MS) and discusses their contribution to several areas of pharmaceutical research: the automation of MS for high-throughput analysis to support new entity research, the use of liquid chromatography (LC)-MS for mixture analysis of degradation products and drug metabolites, the expanded role of highly sensitive MS for the structure elucidation of unknown organic compounds (especially natural products), the study of peptides and proteins, and the detection of non-covalent complexes.