Early mobilization in coronavirus-19 patients treated with extracorporeal membrane oxygenation.

BACKGROUND: Coronavirus disease 2019 (COVID-19) pneumonia can be associated with refractory respiratory failure requiring extracorporeal membrane oxygenation(ECMO). Although ECMO has helped many COVID patients, optimal management strategies for these patients remain unknown. METHODS: We conducted a retrospective review of all COVID patients requiring ECMO at our hospital. Six months into the pandemic, we changed our management strategy to focus on early mobilization. The early mobilization effort included tracheostomy within 48 h of cannulation, decreasing sedation, and an aggressive physical and occupational therapy program progressing toward early ambulation while on ECMO. The primary outcome measured was survival to discharge. The primary stratification was based on the mobilization strategy. RESULTS: From 2020 to 2021, 47 COVID patients have been supported with ECMO at our institution. Five are still in the hospital on ECMO. 39 (83%) were supported with venovenous ECMO while 8 (17%) were supported with venoarterial or a right ventricular assist device type configuration. All 47 (100%) were cannulated at bedside with transesophageal echocardiographic guidance. The average age was 47 ± 9 years; 36(77%) were male; and 20 (43%) were Hispanic. The median duration of support was 22 (11-44) days. Excluding those who remain in the hospital and on support, overall survival to discharge was 24/42 (57%). When stratified by mobilization strategy, early tracheostomy and mobilization were associated with significantly improved survival (74% [17/23] vs. 37% [7/19], p = .02). There were no changes in patient acuity or duration of support throughout the study period. CONCLUSION: In conclusion, early tracheostomy, decreased sedation, and aggressive mobilization of COVID-19 ECMO patients is associated with improved survival.

An Interprofessional Approach to Mobilizing Patients With COVID-19 Receiving Extracorporeal Membrane Oxygenation.

OBJECTIVE: To assess survival outcomes with the intervention of an interprofessional mobilization program for patients with COVID-19 who were receiving venovenous extracorporeal membrane oxygenation (VV-ECMO). DESIGN: Preintervention and postintervention retrospective cohort study. METHODS: Survival outcomes of nonmobilized, adult patients (n = 16) with COVID-19 who were receiving VV-ECMO (May 2020 through December 2020) were compared with those of 26 patients who received a mobility care plan (January 2021 through November 2021). In the preintervention group, full sedation and paralysis were used. In the postintervention group, an early mobilization strategy involving interprofessional collaboration was introduced. RESULTS: The postintervention group had improved survival (73.1% vs 43.8%; P < .04); fewer days of receiving paralytics, fentanyl, and midazolam (P < .01 for all); but more days of dexmedetomidine, morphine, and ketamine administration (P < .01 for all). Concomitantly, more patients in the postintervention cohort received oral or transdermal analgesics, oral anxiolytics, and oral antipsychotics (P < .01 for all), and also required more VV-ECMO cannula adjustments (P = .03). CONCLUSION: Early mobilization of patients with COVID-19 who were receiving VV-ECMO improved survival rates but led to more cannula adjustments.

Blood Product Utilization in Patients With COVID-19 on ECMO.

INTRODUCTION: Although extracorporeal membrane oxygenation (ECMO) has been associated with improved outcomes in COVID patients with respiratory failure, data regarding the need for blood product utilization in this population is inadequate. METHODS: We conducted a retrospective review of all COVID patients requiring ECMO support at our facility. Patient demographics, co-morbidities, measures of acuity, and blood product utilization were identified. Patients were stratified by the presence of a major bleed and the need for dialysis. The primary outcome was blood product utilization. Linear regression models were used to assess predictors of the need for blood products. RESULTS: From 2020 to 2021, 41 patients with COVID-19 were included in our study. Overall 1601 d of support, COVID ECMO patients received 755 units of packed red blood cells (PRBC), 51 units of fresh frozen plasma (FFP), 326 platelets, and 1702 cryoprecipitate, amounting to 18.4 units PRBC per patient or 3.30 units per week of ECMO support. Both major bleeding and the need for dialysis were associated with higher rates of transfusion of PRBC, FFP, and platelets. The average non-bleeding COVID ECMO patient who did not need dialysis required 2.17 units of PRBC, 0.12 units of FFP, 0.76 platelets, and 8.36 of cryoprecipitate per week of ECMO support. On multivariable linear regression analysis, each day on ECMO was associated with 0.30 [0.19-0.42, P < 0.01] units of PRBC. CONCLUSIONS: In conclusion, COVID ECMO is associated with a significant need for blood and blood products. Major bleeding and dialysis are important drivers of blood product requirements.

Intermediate-term survival and functional outcomes of COVID-19 extracorporeal membrane oxygenation patients.

BACKGROUND: Extracorporeal membrane oxygenation (ECMO) has been associated with acceptable short-term survival in patients with refractory respiratory failure secondary to coronavirus disease 2019 (COVID-19) pneumonia. Previous studies have demonstrated acceptable long-term outcomes in patients supported with ECMO for respiratory failure of other etiologies. However, long-term survival and functional outcomes in COVID ECMO patients remain unknown. METHODS: We conducted a retrospective review of all COVID patients requiring ECMO at our hospital. The primary outcomes measured were survival to discharge and contemporary survival. Secondary outcomes included two simple functional assessments: the ongoing need for oxygen supplementation and the ability to return to work. Survival was calculated using the Kaplan-Meier method. Hazard ratios were calculated using Cox hazards regression models. RESULTS: From 2020 to 2021, 48 COVID patients have been supported with ECMO at our hospital. Four patients remain on support and were excluded from further analysis. The average age was 47 ± 8 years, 34 (77%) were males, and the plurality (19, 43%) were Hispanic. Median duration of support was 23 (12-51) days. Median follow-up was 106 (29-226) days. Survival to discharge was 59%. Kaplan-Meier 180-day survival was 51%. Long-term survival conditioned on survival to discharge was 89%. In evaluating functional outcomes, the overwhelming majority of patients no longer required oxygen supplementation (74%), and most had returned to work (52%). CONCLUSION: In conclusion, COVID ECMO patients have acceptable intermediate-term survival with adequate functional recovery.

Sample Mass Estimate for the Use of Near-Infrared and Raman Spectroscopy to Monitor Content Uniformity in a Tablet Press Feed Frame of a Drug Product Continuous Manufacturing Process.

Recently, feed frame-based process analytical technology measurements used to assure product quality during continuous manufacturing processes have received significant attention. These measurements are able to accurately determine uniformity of the powder blend before compression, and in these applications, it is necessary to understand the interrogated sample volume per measurement. This understanding ensures that the blend measurement can be indicative of the uniformity of the final dosage form. A scientifically sound approach is proposed here to estimate sample mass for a continuous manufacturing process that utilizes either near infrared or Raman spectroscopy. A wide range of commercially available probes with varying spot diameters are considered. By comparing near infrared and Raman spectroscopy, an optimal range of probe spot diameters was identified in order to reach an estimated sample mass between 50 and 500 mg for pharmaceutical blends per measurement, which is equivalent to common tablet weight ranges for solid oral dosage forms currently on the market.

Characterization of Near-Infrared and Raman Spectroscopy for In-Line Monitoring of a Low-Drug Load Formulation in a Continuous Manufacturing Process.

Reflectance spectroscopy is an excellent candidate for process analytical technology (PAT) applications in continuous manufacturing of pharmaceutical tablets. Spectroscopic methods provide a real-time, nondestructive measurement of the active pharmaceutical ingredient (API) concentration in order to ensure product quality and uniformity. Of particular challenge is the powder blends with low drug loads (<5% w/w) where the measurement of the signal-to-noise and, in turn, precision limit the ability of the method. We evaluate both near-infrared (NIR) and Raman spectroscopy for use in PAT applications by measuring pharmaceutical blends of varying active ingredient concentrations. Both spectrometers are equipped with a fiber-optically coupled probe head for noncontact measurement of powder blends. A mockup of the interface between the spectrometer and powders within the feed frame of a rotary tablet press is used to simulate the movement of powder blends from the mixer to the press. A port on the feed frame allows measurement by NIR or Raman spectroscopy of the blends just before tablet compression. For our model compound, Raman spectroscopy is shown to have a lower limit-of-detection and less day-to-day variability than NIR spectroscopy. Raman spectroscopy was chosen as the PAT platform to support process development, and working distance and spot size were both optimized for use in the feed-frame of a tablet press. Sufficient limit-of-detection was achieved for monitoring active pharmaceutical ingredient concentrations (API) down to 1% w/w during a semicontinuous manufacturing of tablets. An innovative optimization-based model (EIOT) was used to trend API concentration and demonstrated that the process could be capable of detecting out-of-trend material.

Development of Near Infrared Spectroscopy-based Process Monitoring Methodology for Pharmaceutical Continuous Manufacturing Using an Offline Calibration Approach.

A near-infrared (NIR) calibration was developed using an efficient offline approach to enable a quantitative partial least-squares (PLS) chemometric model to measure and monitor the concentration of active pharmaceutical ingredients (API) in powder blends in the feed frame (FF) of a tablet press. The approach leveraged an offline "feed frame table," which was designed to mimic the full process from a NIR measurement perspective, thereby facilitating a more robust model by allowing more sources of variability to be included in the calibration by minimizing the consumption of API and other raw materials. The design of experiment (DOE) for the calibration was established by an initial risk assessment and included anticipated variability from factors related to formulation, process, environment, and instrumentation. A test set collected on the feed frame table was used to refine the PLS model. Additional fully independent test sets collected from the continuous drug product manufacturing process not only demonstrated the accuracy and precision of the model but also illustrated its robustness to material variability and process variability including mass flow rate and feed frame paddle speed. Further, it demonstrated that a calibration can be generated on the offline feed frame table and then successfully implemented on the full process equipment in a robust manner. Additional benefits of using the feed frame table include streamline model monitoring and maintenance activities in a manufacturing setting. The real-time monitoring enabled by this offline calibration approach can be useful as a key component of the control strategy for continuous manufacturing processes for drug products, including detecting special cause variations such as transient disturbances and enabling product collection/rejection based upon predetermined concentration limits, and may play an important role in enabling real-time release testing (RTRt) for manufactured pharmaceutical products.

Kilogram-scale prexasertib monolactate monohydrate synthesis under continuous-flow CGMP conditions.

Advances in drug potency and tailored therapeutics are promoting pharmaceutical manufacturing to transition from a traditional batch paradigm to more flexible continuous processing. Here we report the development of a multistep continuous-flow CGMP (current good manufacturing practices) process that produced 24 kilograms of prexasertib monolactate monohydrate suitable for use in human clinical trials. Eight continuous unit operations were conducted to produce the target at roughly 3 kilograms per day using small continuous reactors, extractors, evaporators, crystallizers, and filters in laboratory fume hoods. Success was enabled by advances in chemistry, engineering, analytical science, process modeling, and equipment design. Substantial technical and business drivers were identified, which merited the continuous process. The continuous process afforded improved performance and safety relative to batch processes and also improved containment of a highly potent compound.

Investigation of the mechanism of racemization of litronesib in aqueous solution: unexpected base-catalyzed inversion of a fully substituted carbon chiral center.

Mitosis inhibitor (R)-litronesib (LY2523355) is a 1,3,4-thiadiazoline-bearing phenyl and N-(2-ethylamino)ethanesulfonamido-methyl substituents on tetrahedral C5. Chiral instability has been observed at pH 6 and above with the rate of racemization increasing with pH. A positively charged trigonal intermediate is inferred from the fact that p-methoxy substituent on the phenyl accelerated racemization, whereas a p-trifluoromethyl substituent had the opposite effect. Racemization is proposed to occur through a relay mechanism involving intramolecular deprotonation of the sulfonamide by the side chain amino group and attack of the sulfonamide anion on C5, cleaving the C5S bond, to form an aziridine; heterolytic dissociation of the aziridine yields an ylide. This pathway is supported by (1) a crystal structure providing evidence for a hydrogen bond between the sulfonamide NH and the amino group, (2) effects of substituents on the rate of racemization, and (3) computational studies. This racemization mechanism results from neighboring group effects in this densely functionalized molecule. Of particular novelty is the involvement of the side-chain secondary amino group, which overcomes the weak acidity of the sulfonamide by anchimeric assistance.

On-column nitrosation of amines observed in liquid chromatography impurity separations employing ammonium hydroxide and acetonitrile as mobile phase.

The availability of high performance liquid chromatography (HPLC) columns capable of operation at pH values up to 12 has allowed a greater selectivity space to be explored for method development in pharmaceutical analysis. Ammonium hydroxide is of particular value in the mobile phase because it is compatible with direct interfacing to electrospray mass spectrometers. This paper reports an unexpected N-nitrosation reaction that occurs with analytes containing primary and secondary amines when ammonium hydroxide is used to achieve the high pH and acetonitrile is used as the organic modifier. The nitrosation reaction has generality. It has been observed on multiple columns from different vendors and with multiple amine-containing analytes. Ammonia was established to be the source of the nitroso nitrogen. The stainless steel column frit and metal ablated from the frit have been shown to be the sites of the reactions. The process is initiated by removal of the chromium oxide protective film from the stainless steel by acetonitrile. It is hypothesized that the highly active, freshly exposed metals catalyze room temperature oxidation of ammonia to NO but that the actual nitrosating agent is likely N(2)O(3).

Civil commitment for substance use disorder patients under the Florida Marchman Act: demographics and outcomes in the private clinical setting.

The Florida Marchman Act, a statutory process for civil commitment of persons with substance use disorders. The paper describes the various methods by which the Act may be employed, and examines the demographics and outcomes of 100 patients admitted to a private treatment setting pursuant to Marchman Act authority.

A systematic approach to development of liquid chromatographic impurity methods for pharmaceutical analysis.

A strategy for developing chromatographic methods designed to determine impurities and degradation products in active pharmaceutical ingredients and drug products is presented. Selectivity is achieved by evaluating a chromatographic space comprised of 12 stationary/mobile phase combinations. Stationary phases predicted to be orthogonal based on their hydrophobic subtraction model parameters used. The particle sizes, column dimensions, and gradient times chosen provide high peak capacities and allow operation at backpressures that can be achieved with standard instrumentation. The mobile phases utilized are compatible with MS detection and cover a wide range of pH, solvent strength, and solvent selectivity. Analyte detection is accomplished using a combination of diode array and mass spectroscopic detectors which allow mixtures of project compounds to be injected and selectively detected. Automation of data acquisition and processing is accomplished using AutoChrom software from ACD\Labs. The strategy is illustrated with detailed data from two case studies and summary data from nineteen pharmaceutical projects.

Mechanotransduction and anoikis: death and the homeless cell.

Developed organs display strict spatial organization of differentiated cells which is required for proper organ function. One important device that prevents tissue disorganization is the death of cells that lose anchorage to their native matrix, a signal that indicates potential loss of proper tissue context. Termed anoikis (Greek for Homelessness), this form of cell death is a specialized form of apoptosis. Interestingly, at certain stages of development and tissue repair, cells are required to migrate in an unanchored state, suggesting that anoikis must be strictly regulated at some level. Likewise, cellular transformation is often accompanied by an inappropriate loss of anoikis and subsequent acquisition of a metastatic phenotype. Despite its importance, the molecular pathways involved in the regulation of anoikis and the proximal signals reporting loss of anchorage are poorly understood. Recent studies suggest that attachment may be reported by a mechanosensory testing of the cell's physical environment.

An analysis of the results from Two-Generation Reproduction Toxicity Studies to assess the value of mating animals of the second (F1) generation for the detection of adverse treatment-related effects on reproductive performance.

The need to routinely perform a two-generation study to assess reproductive performance is under debate. This review has analysed the results from 22 consecutive Two-Generation Reproduction Toxicity Studies performed within our laboratories, to determine the value of mating each generation in the detection of treatment-related effects on reproductive performance. The chemicals included 13 agrochemicals, 5 industrial chemicals and 4 food additives. Four chemicals (all from the agrochemical group) were found to have clear treatment-related effects and in three studies effects were confined to the second (F1) generation. For the three studies with second (F1) generation effects, we applied the triggers, proposed by the Agricultural Chemical Safety Assessment (ASCA) Technical Committee of HESI, for extension of the one-generation study and found that mating of the second (F1) generation would have been triggered in these studies by effects on the F1 offspring growth and sexual maturation. No effects were seen in the original parental (F0) generation which would have activated the change from a one- to a two-generation study.

HIV-1 Tat activates dual Nox pathways leading to independent activation of ERK and JNK MAP kinases.

Human immunodeficiency virus, type 1 Tat is known to exert pleiotropic effects on the vascular endothelium through mitogen-activated protein (MAP) kinases, although the signaling pathways leading to MAP kinase activation are incompletely understood. We focused on proximal pathways potentially governing downstream MAP kinase activity by Tat. Within 2 min, Tat activated both Ras and Rho GTPases in endothelial cells, leading to ERK phosphorylation by 10 min. Notably, Rac1 was necessary for downstream activation of RhoA and both Rac1 and RhoA acted upstream of the Ras/ERK cassette. Antioxidants and the oxidase inhibitor diphenylene iodonium blocked ERK phosphorylation, but specific interference with the canonical Nox2 oxidase had no effect on ERK. Instead, knock down of the novel oxidase Nox4 completely suppressed Tat-dependent Ras and ERK activation downstream of Rac1 and RhoA. Conversely, interference with Rac1, PAK1, and Nox2 blocked JNK phosphorylation, whereas RhoA(N19) and Nox4 knock down did not. Further, knock down of Nox2, but not Nox4, blocked Tat-induced cytoskeletal rearrangement, whereas knock down of Nox4, but not Nox2, blocked Tat-dependent proliferation. Rac1, therefore, bifurcates Tat signaling, leading to concurrent but separate Nox4-dependent Ras/ERK activation, and Nox2-dependent JNK activation. Tat signaling, therefore, provides an example of Nox-specific differential control of MAP kinase pathways.

p66Shc mediates anoikis through RhoA.

Detachment of parenchymal cells from a solid matrix switches contextual cues from survival to death during anoikis. Marked shape changes accompany detachment and are thought to trigger cell death, although a working model to explain the coordination of attachment sensation, shape change, and cell fate is elusive. The constitutive form of the adapter Shc, p52Shc, confers survival properties, whereas the longer p66Shc signals death through association with cytochrome c. We find that cells that lack p66Shc display poorly formed focal adhesions and escape anoikis. However, reexpression of p66Shc restores anoikis through a mechanism requiring focal adhesion targeting and RhoA activation but not an intact cytochrome c-binding motif. This pathway stimulates the formation of focal adhesions and stress fibers in attached cells and tension-dependent cell death upon detachment. p66Shc may thus report attachment status to the cell by imposing a tension test across candidate anchorage points, with load failure indicating detachment.

Treatment for attorneys with substance related and co-occurring psychiatric disorders: demographics and outcomes.

The following paper is the result of a retrospective study of clinical case files of legal professionals treated for substance-related and co-occurring psychiatric disorders at a recovery center specializing in the care of impaired professionals. Attorneys traditionally differ from healthcare professionals in two important ways: first, they prematurely leave treatment in greater numbers, and second, they suffer a higher incidence of co-occurring psychiatric disorders. Sixty percent of the attorneys presented with concurrent psychiatric conditions (Axis I and Axis II), compared to 46% of their healthcare colleagues. More than half of the lawyers treated had a prior history of criminal arrest. Sixty-four percent completed treatment compared to an 86% completion rate for medical professionals. The completion rate for lawyers improved significantly following institution of a dedicated impaired attorneys' program under the direction of an attorney/clinician in October of 1999. Prospective studies are needed to determine if attorneys are at greater risk of developing substance related and/or psychiatric disorders than are other professionals of similar demographic backgrounds and what specialized intervention, treatment, and case management services might be necessary to assure equivalent outcomes.

Ornithine decarboxylase promotes catalysis by binding the carboxylate in a buried pocket containing phenylalanine 397.

Ornithine decarboxylase (ODC) is a pyridoxal 5'-phosphate (PLP) dependent enzyme that catalyzes the decarboxylation of l-Orn to putrescine, a rate-limiting step in the formation of polyamines. The X-ray crystal structures of ODC, complexed to several ligands, support a model where the substrate is oriented with the carboxyl-leaving group buried on the re face of the PLP cofactor. This binding site is composed of hydrophobic and electron-rich residues, in which Phe-397 is predicted to form a close contact. Mutation of Phe-397 to Ala reduces the steady-state rate of product formation by 150-fold. Moreover, single turnover analysis demonstrates that the rate of the decarboxylation step is decreased by 2100-fold, causing this step to replace product release as the rate-limiting step in the mutant enzyme. These data support the structural prediction that the carboxyl-leaving group is positioned to interact with Phe-397. Multiwavelength stopped-flow analysis of reaction intermediates suggests that a major product of the reaction with the mutant enzyme is pyridoximine 5'-phosphate (PMP), resulting from incorrect protonation of the decarboxylated intermediate at the C4' position. This finding was confirmed by HPLC analysis of the reaction products, demonstrating that Phe-397 also plays a role in maintaining the integrity of the reaction chemistry. The finding that the carboxylate-leaving group is oriented on the buried side of the PLP cofactor suggests that ODC facilitates decarboxylation by destabilizing the charged substrate carboxyl group in favor of an electrostatically more neutral transition state.