Pharmacogenomics of GPCR Drug Targets.

Natural genetic variation in the human genome is a cause of individual differences in responses to medications and is an underappreciated burden on public health. Although 108 G-protein-coupled receptors (GPCRs) are the targets of 475 (∼34%) Food and Drug Administration (FDA)-approved drugs and account for a global sales volume of over 180 billion US dollars annually, the prevalence of genetic variation among GPCRs targeted by drugs is unknown. By analyzing data from 68,496 individuals, we find that GPCRs targeted by drugs show genetic variation within functional regions such as drug- and effector-binding sites in the human population. We experimentally show that certain variants of µ-opioid and Cholecystokinin-A receptors could lead to altered or adverse drug response. By analyzing UK National Health Service drug prescription and sales data, we suggest that characterizing GPCR variants could increase prescription precision, improving patients' quality of life, and relieve the economic and societal burden due to variable drug responsiveness. VIDEO ABSTRACT.

Comprehensive assessment of TECENTRIQ® and OPDIVO®: analyzing immunotherapy indications withdrawn in triple-negative breast cancer and hepatocellular carcinoma.

Atezolizumab (TECENTRIQ®) and nivolumab (OPDIVO®) are both immunotherapeutic indications targeting programmed cell death 1 ligand 1 (PD-L1) and programmed cell death 1 (PD-1), respectively. These inhibitors hold promise as therapies for triple-negative breast cancer (TNBC) and hepatocellular carcinoma (HCC) and have demonstrated encouraging results in reducing the progression and spread of tumors. However, due to their adverse effects and low response rates, the US Food and Drug Administration (FDA) has withdrawn the approval of atezolizumab in TNBC and nivolumab in HCC treatment. The withdrawals of atezolizumab and nivolumab have raised concerns regarding their effectiveness and the ability to predict treatment responses. Therefore, the current study aims to investigate the immunotherapy withdrawal of PD-1/PD-L1 inhibitors, specifically atezolizumab for TNBC and nivolumab for HCC. This study will examine both the structural and clinical aspects. This review provides detailed insights into the structure of the PD-1 receptor and its ligands, the interactions between PD-1 and PD-L1, and their interactions with the withdrawn antibodies (atezolizumab and nivolumab) as well as PD-1 and PD-L1 modifications. In addition, this review further assesses these antibodies in the context of TNBC and HCC. It seeks to elucidate the factors that contribute to diverse responses to PD-1/PD-L1 therapy in different types of cancer and propose approaches for predicting responses, mitigating the potential risks linked to therapy withdrawals, and optimizing patient outcomes. By better understanding the mechanisms underlying responses to PD-1/PD-L1 therapy and developing strategies to predict these responses, it is possible to create more efficient treatments for TNBC and HCC.

Regulation of Molecular Diagnostics.

Molecular diagnostic tests enable rapid analysis of genomic and proteomic markers. These tests are subject to diverging premarket access and postmarket surveillance requirements and mechanisms in the United States and the European Union. Each of these jurisdictions has its own challenges in keeping the regulations up to date with technological developments. A specific area of attention is that of laboratory-developed tests in the United States and health institution in-house-produced tests in the European Union, for which the United States and the European Union have markedly different regulatory approaches. Both jurisdictions have specific but differing requirements for the use of test samples and test-related data under their rules regarding the protection of (personal) health data, which can cause complexity when moving samples or sample-related data from one jurisdiction to the other.

A practical guide for implementing omalizumab therapy for food allergy.

The recent approval of omalizumab for the treatment of IgE-mediated food allergy is an important step forward for the millions of food allergy patients in the United States. Through the depletion of circulating IgE and the subsequent reduction of FcεR1 on key effector cells, patients increase their tolerance to food allergens. However, omalizumab does not permit patients to eat foods that they are allergic to with impunity. Rather, it protects them from most accidental exposures. In addition, omalizumab does not cure food allergy and has not demonstrated true immunomodulation. Thus, omalizumab might be a lifelong therapy for some patients. Furthermore, there are many important questions and issues surrounding the appropriate administration of omalizumab to treat food allergy, which we discuss. Managing treatment of patients with disease that falls outside the dosing range, assessing treatment response or nonresponse, addressing its appropriateness for patients older than 55, and determining whether immunotherapy plus omalizumab provides any advantage over omalizumab alone all need to be examined. Identifying appropriate patients for this therapy is critical given the cost of biologics. Indeed, not all food allergy patients are good candidates for this therapy. Also, when and how to stop omalizumab therapy in patients who may have outgrown their food allergy needs to be elucidated. Thus, although this therapy provides a good option for patients with food allergies, much information is needed to determine how best to use this therapy. Despite many unanswered questions and issues, we provide clinicians with some practical guidance on implementing this therapy in their patients.

Escape Velocity-the Launch of Microbiome Therapies.

Food and Drug Administration approval of the first microbiome therapies represents a true expansion the treatment paradigm for Clostridioides difficile but raises new questions about the future role of fecal microbiota transplantation. The authors outline the advances in microbiome therapeutic development that have addressed fecal microbiota transplantation's (FMT's) inherent limitations of safety and scalability. The authors also suggest that as microbiome therapeutic development continues for other indications, FMT will likely remain a necessary model of human microbiota dynamics for translational research.

Benzopyrone, a privileged scaffold in drug discovery: An overview of FDA-approved drugs and clinical candidates.

Natural products have always served as an important source of drugs for treating various diseases. Among various privileged natural product scaffolds, the benzopyrone class of compounds has a substantial presence among biologically active compounds. One of the pioneering anticoagulant drugs, warfarin approved in 1954 bears a benzo-α-pyrone (coumarin) nucleus. The widely investigated psoriasis drugs, methoxsalen, and trioxsalen, also contain a benzo-α-pyrone nucleus. Benzo-γ-pyrone (chromone) containing drugs, cromoglic acid, and pranlukast were approved as treatments for asthma in 1982 and 2007, respectively. Numerous other small molecules with a benzopyrone core are under clinical investigation. The present review discusses the discovery, absorption, distribution, metabolism, excretion properties, and synthetic approaches for the Food and Drug Administration-approved and clinical-stage benzopyrone class of compounds. The role of the pyrone core in biological activity has also been discussed. The present review unravels the potential of benzopyrone core in medicinal chemistry and drug development.

Babesia BdFE1 esterase is required for the anti-parasitic activity of the ACE inhibitor fosinopril.

Effective and safe therapies for the treatment of diseases caused by intraerythrocytic parasites are impeded by the rapid emergence of drug resistance and the lack of novel drug targets. One such disease is human babesiosis, which is a rapidly emerging tick-borne illness caused by Babesia parasites. In this study, we identified fosinopril, a phosphonate-containing, FDA-approved angiotensin converting enzyme (ACE) inhibitor commonly used as a prodrug for hypertension and heart failure, as a potent inhibitor of Babesia duncani parasite development within human erythrocytes. Cell biological and mass spectrometry analyses revealed that the conversion of fosinopril to its active diacid molecule, fosinoprilat, is essential for its antiparasitic activity. We show that this conversion is mediated by a parasite-encoded esterase, BdFE1, which is highly conserved among apicomplexan parasites. Parasites carrying the L238H mutation in the active site of BdFE1 failed to convert the prodrug to its active moiety and became resistant to the drug. Our data set the stage for the development of this class of drugs for the therapy of vector-borne parasitic diseases.

Guidance for securing approvals for new biomarkers: from discovery to clinical implementation.

The journey of translating a molecular discovery into the clinic involves multiple steps and requires planning, time, effort, and money. In this review, we provide a quick guide on the technical and clinical validation parameters that are necessary for successful commercialization of molecular and other markers. We also briefly address the different options for regulatory approvals. Successful clinical implantation depends on rigorous technical and clinical validation, and the ability to develop clear guidelines for the indications for testing (i.e. which patients are eligible to have this test), the frequency of testing, and also a clear interpretation of test results. Successful implementation requires providing evidence that the results of this test can be used to improve patient care. There are currently multiple routes for implementation of clinical molecular tests, which include regulatory agency- approved companion diagnostics, laboratory developed tests, or direct-to-consumer testing. Regulatory approval is considered the gold-standard, but it requires time and resources. There is an ongoing debate about the need for regulatory approval of laboratory developed testing. Ongoing oversight is maintained through lab accreditation and proficiency testing programs, which provide a common approach to ensuring high standards and consistent performance in clinical molecular labs. Before moving into the clinic, confirmation of both the clinical and analytic validity of a new molecular test is essential.

Recapitulation of human pathophysiology and identification of forensic biomarkers in a translational model of chlorine inhalation injury.

Chlorine gas (Cl2) has been repeatedly used as a chemical weapon, first in World War I and most recently in Syria. Life-threatening Cl2 exposures frequently occur in domestic and occupational environments, and in transportation accidents. Modeling the human etiology of Cl2-induced acute lung injury (ALI), forensic biomarkers, and targeted countermeasures development have been hampered by inadequate large animal models. The objective of this study was to develop a translational model of Cl2-induced ALI in swine to understand toxico-pathophysiology and evaluate whether it is suitable for screening potential medical countermeasures and to identify biomarkers useful for forensic analysis. Specific pathogen-free Yorkshire swine (30-40 kg) of either sex were exposed to Cl2 (≤240 ppm for 1 h) or filtered air under anesthesia and controlled mechanical ventilation. Exposure to Cl2 resulted in severe hypoxia and hypoxemia, increased airway resistance and peak inspiratory pressure, and decreased dynamic lung compliance. Cl2 exposure resulted in increased total leucocyte and neutrophil counts in bronchoalveolar lavage fluid, vascular leakage, and pulmonary edema compared with the air-exposed group. The model recapitulated all three key histopathological features of human ALI, such as neutrophilic alveolitis, deposition of hyaline membranes, and formation of microthrombi. Free and lipid-bound 2-chlorofatty acids and chlorotyrosine-modified proteins (3-chloro-l-tyrosine and 3,5-dichloro-l-tyrosine) were detected in plasma and lung tissue after Cl2 exposure. In this study, we developed a translational swine model that recapitulates key features of human Cl2 inhalation injury and is suitable for testing medical countermeasures, and validated chlorinated fatty acids and protein adducts as biomarkers of Cl2 inhalation.NEW & NOTEWORTHY We established a swine model of chlorine gas-induced acute lung injury that exhibits several features of human acute lung injury and is suitable for screening potential medical countermeasures. We validated chlorinated fatty acids and protein adducts in plasma and lung samples as forensic biomarkers of chlorine inhalation.

Representation of women in clinical trials supporting FDA-approval of contemporary cancer therapies.

Contemporary anticancer therapies frequently have different efficacy and side effects in men and women. Yet, whether women are well-represented in pivotal trials supporting contemporary anticancer drugs is unknown. Leveraging the Drugs@FDA database, clinicaltrials.gov, MEDLINE, and publicly available FDA-drug-reviews, we identified all pivotal (phase II and III) non-sex specific trials supporting FDA-approval of anticancer drugs (1998-2018). Observed-enrollment-rates were compared to expected-population-rates derived from concurrent US-National-Cancer-Institute's Surveillance-Epidemiology-and-End-Results (SEER) reported rates and US-Census databases. Primary outcome was the proportional representation of women across trials, evaluated by a participation-to-prevalence ratio (PPR), according to cancer type. Secondary outcome was the report of any sex-specific analysis of efficacy and/or safety, irrespective of treatment-arm. Overall, there were 148 trials, enrolling 60,216 participants (60.5 ± 4.0 years, 40.7% female, 79.1% biologic, targeted, or immune-based therapies) evaluating 99 drugs. Sex was reported in 146 (98.6%) trials, wherein 40.7% (24,538) were women, compared to 59.3% (35,678) men (p < .01). Altogether, women were under-represented in 66.9% trials compared to the proportional incidence of cancers by respective disease type; weight-average PPR of 0.91 (relative difference: -9.1%, p < .01). Women were most under-represented in gastric (PPR = 0.63), liver (PPR = 0.71), and lung (PPR = .81) cancer trials. Sex-based safety data was reported in 4.0% trials. There was no association between adequate female enrollment and drug efficacy (HR: 0.616 vs. 0.613, p = .96). Over time, there was no difference in the percentage of women recruited into clinical trials. Among pivotal clinical trials supporting contemporary FDA-approved cancer drugs, women were frequently under-represented and sex-specific-efficacy and safety-outcomes were commonly not reported.

Why partial heart transplantation could be regulated as organ transplantation.

Partial heart transplant (PHT) is a recent clinical innovation involving the transplantation of a segment of the heart (valves) directly from the deceased donor into the recipient patient. This procedure holds out the possibility of significant benefit, especially for pediatric patients because these grafts show growth potential after transplant, reducing or eliminating the current need for repeat procedures. The clinical process for donation and transplant of partial heart (PH) grafts generally follows an organ clinical pathway; however, the Food and Drug Administration has recently stated its intent to regulate PH as tissues, raising a host of regulatory considerations. PHT requires donor testing and eligibility determinations within a short, clinically viable timeframe and, similar to organ transplant, involves donor-recipient matching. Waitlist allocation policies that are a regulatory focus of the Organ Procurement and Transplantation Network including equity and efficiency may become relevant. Oversight of PHT by the Organ Procurement and Transplantation Network could be accomplished through interpretation of the vascular composite allograft definition or through designation by the US Department of Health and Human Services of PH grafts as organs. While some clinical questions remain unanswered, it is important to carefully address these regulatory considerations to support the emergence of this innovation and ensure the continued trust of the donating public and the patients who may benefit from PHT.

Efficacy of therapeutically administered gepotidacin in a rabbit model of inhalational anthrax.

Bacillus anthracis is a Gram-positive Centers for Disease Control and Prevention category "A" biothreat pathogen. Without early treatment, inhalation of anthrax spores with progression to inhalational anthrax disease is associated with high fatality rates. Gepotidacin is a novel first-in-class triazaacenaphthylene antibiotic that inhibits bacterial DNA replication by a distinct mechanism of action and is being evaluated for use against biothreat and conventional pathogens. Gepotidacin selectively inhibits bacterial DNA replication via a unique binding mode and has in vitro activity against a collection of B. anthracis isolates including antibacterial-resistant strains, with the MIC90 ranging from 0.5 to 1 µg/mL. In vivo activity of gepotidacin was also evaluated in the New Zealand White rabbit model of inhalational anthrax. The primary endpoint was survival, with survival duration and bacterial clearance as secondary endpoints. The trigger for treatment was the presence of anthrax protective antigen in serum. New Zealand White rabbits were dosed intravenously for 5 days with saline or gepotidacin at 114 mg/kg/d to simulate a dosing regimen of 1,000 mg intravenous (i.v.) three times a day (TID) in humans. Gepotidacin provided a survival benefit compared to saline control, with 91% survival (P-value: 0.0001). All control animals succumbed to anthrax and were found to be blood- and organ culture-positive for B. anthracis. The novel mode of action, in vitro microbiology, preclinical safety, and animal model efficacy data, which were generated in line with Food and Drug Administration Animal Rule, support gepotidacin as a potential treatment for anthrax in an emergency biothreat situation.

Expert Panel Recommendations: Practical Clinical Applications for Initiating and Monitoring Resmetirom in Patients with MASH/NASH and Moderate to Noncirrhotic Advanced Fibrosis.

Metabolic dysfunction-associated steatotic liver disease affects 1 in 4 people in the United States and western Europe, with an important proportion developing metabolic dysfunction-associated steatohepatitis (MASH), the progressive subtype of metabolic dysfunction-associated steatotic liver disease. Cirrhosis caused by MASH is a leading indication for liver transplantation and the most common cause of hepatocellular carcinoma. Hitherto, there have been no specific pharmacotherapies for MASH. The recent conditional approval by the Food and Drug Administration of resmetirom for the treatment of moderate or advanced MASH presents a much-anticipated therapeutic option for patients with noncirrhotic advanced MASH. Specifically, the intended population for resmetirom are patients with MASH and fibrosis stages 2 or 3. The approval of resmetirom also presents important challenges, including how to noninvasively identify patients with fibrosis stages 2-3, and how to exclude patients with more advanced disease who should not be treated until further data emerge on the use of resmetirom in this population. Herein we consider the available literature with regard to identifying the intended population for treatment with resmetirom and in proposing criteria for stopping treatment.

Applicability of selected reaction monitoring for precise screening tests.

INTRODUCTION: The proactive identification of diseases through screening tests has long been endorsed as a means to preempt symptomatic onset. However, such screening endeavors are fraught with complications, such as diagnostic inaccuracies, procedural risks, and patient unease during examinations. These challenges are amplified when screenings for multiple diseases are administered concurrently. Selected Reaction Monitoring (SRM) offers a unique advantage, allowing for the high-throughput quantification of hundreds of analytes with minimal interferences. AREAS COVERED: Our research posits that SRM-based assays, traditionally tailored for single-disease biomarker profiling, can be repurposed for multi-disease screening. This innovative approach has the potential to substantially alleviate time, labor, and cost demands on healthcare systems and patients alike. Nonetheless, there are formidable methodological hurdles to overcome. These include difficulties in detecting low-abundance proteins and the risk of model overfitting due to the multiple functionalities of single proteins across different disease spectrums - issues especially pertinent in blood-based assays where detection sensitivity is constrained. As we move forward, technological strides in sample preparation, online extraction, throughput, and automation are expected to ameliorate these limitations. EXPERT OPINION: The maturation of mass spectrometry's integration into clinical laboratories appears imminent, positioning it as an invaluable asset for delivering highly sensitive, reproducible, and precise diagnostic results.

Community consensus for Heparan sulfate as a biomarker to support accelerated approval in Neuronopathic Mucopolysaccharidoses.

Mucopolysaccharidoses (MPS) disorders are a group of ultra-rare, inherited, lysosomal storage diseases caused by enzyme deficiencies that result in accumulation of glycosaminoglycans (GAGs) in cells throughout the body including the brain, typically leading to early death. Current treatments do not address the progressive cognitive impairment observed in patients with neuronopathic MPS disease. The rarity and clinical heterogeneity of these disorders as well as pre-existing brain disease in clinically diagnosed patients make the development of new therapeutics utilizing a traditional regulatory framework extremely challenging. Children with neuronopathic MPS disorders will likely sustain irreversible brain damage if randomized to a placebo or standard-of-care treatment arm that does not address brain disease. The United States Food and Drug Administration (FDA) recognized these challenges, and, in 2020, issued final guidance for industry on slowly progressive, low-prevalence, rare diseases with substrate deposition that result from single enzyme defects, outlining a path for generating evidence of effectiveness to support accelerated approval based on reduction of substrate accumulation [1]. Neuronopathic MPS disorders, which are characterized by the accumulation of the GAG heparan sulfate (HS) in the brain, fit the intended disease characteristics for which this guidance was written, but to date, this guidance has not yet been applied to any therapeutic candidate for MPS. In February 2024, the Reagan-Udall Foundation for the FDA convened a public workshop for representatives from the FDA, patient advocacy groups, clinical and basic science research, and industry to explore a case study of using cerebrospinal fluid (CSF) HS as a relevant biomarker to support accelerated approval of new therapeutics for neuronopathic MPS disorders. This review provides a summary of the MPS presentations at the workshop and perspective on the path forward for neuronopathic MPS disorders.

Financial conflicts of interest among presenters, panellists and moderators at haematology and oncology FDA workshops.

OBJECTIVE: To assess the characteristics and financial conflicts of interest of presenters, panellists and moderators at haematology and oncology workshops held jointly with or hosted by the US FDA. SETTING: We included information on all publicly available haematology or oncology FDA workshop agendas held between 1 January 2018 and 31 December 2022. EXPOSURE: General and research payments reported on Open Payments, industry funding to patient advocacy organizations reported on their webpages or 990 tax forms and employment in both pharmaceutical and regulatory settings. RESULTS: Among physicians eligible for payments, 78% received at least one payment from the industry between 2017 and 2021. The mean general payment amount was $82,170 for all years ($16,434 per year) and the median was $14,906 for all years ($2981 per year). Sixty-nine per cent of patient advocacy speakers were representing organizations that received financial support from the pharmaceutical industry. Among those representing regulatory agencies or pharmaceutical companies, 16% had worked in both settings during their careers. CONCLUSIONS AND RELEVANCE: Our findings in this cross-sectional study show a majority of US-based physician presenters at haematology and oncology workshops held jointly with members of the US FDA have some financial conflict of interest with the pharmaceutical industry. These findings support the need for clear disclosures and suggest that a more balanced selection of presenters with fewer conflicts may help to limit bias in discussions between multiple stakeholders.

Relationship between Anaplastic Lymphoma Kinase Inhibitors and Epileptic Seizure Disorder: A Post-Marketing Surveillance Study.

INTRODUCTION: Anaplastic lymphoma kinase (ALK) has been to be involved in the uptake and regulation of dopamine 2 receptor (D2R), a G protein-coupled receptor expressed in various brain regions. Therefore, it is crucial to understand the relationship between ALK inhibitors and seizures is an important issue. This study investigated the relationship between ALK inhibitors and seizures. METHODS: This study investigated the relationship between ALK inhibitors and seizures through a disproportionality analysis using the US Food and Drug Administration (FDA) Adverse Event Reporting System (FAERS). The target drugs were the ALK inhibitors crizotinib, ceritinib, alectinib, brigatinib, and lorlatinib. The seizures covered were defined high-level group term (HLGT): "Seizures (incl. subtype)" including high-level term (HLT): "seizures and seizure disorders NEC." This study used the information component (IC), a signal score, as a Bayesian statistical method for disproportionality analysis. The signal detection criteria used in this study were the same as those reported previously: a lower limit of 95% credible interval (CrI) for IC >0. RESULTS: The signal scores of '"seizures and seizure disorders not elsewhere classified (NEC)" "for each ALK inhibitor were crizotinib (IC: -0.00052, 95% CrI: -0.38-0.27), ceritinib (IC: 1.18, 95% CrI: 0.68-1.54), alectinib (IC: 0.68, 95% CrI: 0.19-1.02), brigatinib (IC: 1.04, 95% CrI: 0.32-1.54), and lorlatinib (IC: 0.82, 95% CrI: 0.11-1.32). On the other hand, "generalized tonic-clonic seizures," "partial simple seizures NEC," "absence seizures," and "partial complex seizures" had no or few reported cases, and no signal was detected. CONCLUSION: To our knowledge, this is the first report to evaluate the relationship between ALK inhibitors and seizures using post-marketing surveillance data. These results suggest that ceritinib, alectinib, brigatinib, and lorlatinib, which are highly brain-migrating drugs, are associated with seizures.

Clearance of residual genome editing components used for ex vivo genome-editing of allogeneic cell therapy products.

BACKGROUND AIMS: Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)-associated genome editing (GE) components (e.g., nucleases, guide RNAs (gRNAs), and plasmids) are used to genetically modify cells during development of ex vivo genome-edited cell therapies. Prolonged presence of GE components may increase the risk of unintended genome modifications (e.g., off-target editing and chromosomal rearrangements). This risk is a function of the stability of the GE components, culture conditions (i.e., culture length, media changes, etc.), and the nature of the GE component (i.e., only plasmids can be integrated into a cell's genome). Testing for residual GE components on ex vivo genetically edited drug products is generally recommended in current regulatory guidance (CBER 2024). For allogenic cell therapies derived from induced pluripotent stem cells (iPSC), cells typically undergo clonal selection and extensive culturing following completion of genome editing. This post-engineering clonal selection substantially reduces the amount of residual GE components while the long-duration cell culture significantly reduces the presence of active residual GE components. Here we present a case in which the need for testing of the drug product for residual GE components has been eliminated. METHODS: In silico modeling was used to estimate clearance mechanisms across a variety of relevant assumptions, including disposition of extracellular GE components via media changes and dilution of intracellular GE components via cell expansion. Determining the ability of each GE component-alone or in complex with other GE components-to modify genomic material was assessed by a series of both in vitro and ex vivo (i.e., engineering cells) studies. For the in vitro studies, a DNA cutting assay was developed to assess the ability of the component to cut a representative DNA strand. For the ex vivo modification of cells, an assessment of the knock-out of the relevant gene was completed by flow cytometry specifically assessing the presence or absence of protein expression on the modified cells. The persistence and stability of GE components were examined under cell-mimicking conditions and in ex vivo modified cells. The components were stressed under multiple conditions mimicking a range of culture conditions and tested in the aforementioned DNA cutting assay. The presence of residual gRNA was directly assessed in the ex vivo modified cells via a gRNA-specific digital droplet polymerase chain reaction (ddPCR) assay. RESULTS: Simulations estimating genome editing residual clearance via dilution for extracellular residuals (via media changes) or intracellular residuals (via cell doubling) demonstrate clearance of measurable residuals within 28 days of cell culture. Studies simulating the stability of genome editing residuals estimate less than 7 days for the nuclease, gRNA and ribonucleoprotein (RNP) complex. gRNA was undetectable by 8 days post-engineering under actual engineering conditions. Additionally, without gRNA present, CRISPR Cas12a nucleases did not demonstrate evidence of cutting. In contrast, plasmid DNA can be randomly integrated into the genome and free plasmid is highly stable under cell culture-like conditions (50+ days). Additionally, plasmid DNA integrated in cells will propagate during mitosis, leading to the additional risk of expansion of an unintentional integration event. CONCLUSIONS: Both the gRNA and nuclease in the RNP complex are required for DNA cutting. Neither individual component nor the complex are stable beyond 7 days in culture-mimicking conditions. These findings suggest that the risk of unplanned genomic modification resulting from residual gRNA or nuclease is minimal for processes in which extensive culture is performed after the completion of genome editing and clonal selection. However, the risk of residual plasmid DNA integration is significantly higher regardless of the manufacturing process. The residual plasmid itself is quite stable (at least 50 days) and the risk of random, off-target integration is present. By establishing the stability of these components, we have demonstrated that testing for residual gRNA or nuclease is not warranted for clonally derived allogeneic cell therapies.

Defect-Engineered Metal-Organic Framework/Polyimide Mixed Matrix Membrane for CO2 Separation.

Defect-engineered metal-organic frameworks (MOFs) with outstanding structural and chemical features have become excellent candidates for specific separation applications. The introduction of structural defects in MOFs as an efficient approach to manipulate their functionality provides excellent opportunities for the preparation of MOF-based mixed matrix membranes (MMMs). However, the use of this strategy to adjust the properties and develop the separation performance of gas separation membranes is still in its early stages. Here, a novel defect-engineered MOF (quasi ZrFum or Q-ZrFum) was synthesized via a controlled thermal deligandation process and incorporated into a CO2-philic 6FDA-durene polyimide (PI) matrix to form Q-ZrFum loaded MMMs. Defect-engineered MOFs and fabricated MMMs were investigated regarding their characteristic properties and separation performance. The incorporation of defects into the MOF structure increases the pore size and provides unsaturated active metal sites that positively affect CO2 molecule transport. The interfacial compatibility between the Q-ZrFum particles and the PI matrix increases via the deligandation process, which improves the mechanical strength of Q-ZrFum loaded membranes. MMM containing 5 wt.% of defect-engineered Q-ZrFum exhibits excellent CO2 permeability of 1308 Barrer, which increased by 99 % compared to the pure PI membrane (656 Barrer) at a feed pressure of 2 bar. CO2/CH4 and CO2/N2 selectivity reached 44 and 26.6 which increased by about 70 and 16 %, respectively. This study emphasizes that defect-engineered MOFs can be promising candidates for use as fillers in the preparation of MMMs for the future development of membrane-based gas separation applications.

Chimeric antigen receptor T-cell therapy-induced nervous system toxicity: a real-world study based on the FDA Adverse Event Reporting System database.

BACKGROUND: Nervous system toxicity (NST) is one of the most frequent and dangerous side effects of chimeric antigen receptor T-cell (CAR-T) therapy, which is an effective treatment for related tumors in most relapsed/refractory (r/r) hematologic malignancies. Current clinical trial data do not fully reflect the real-world situation. Therefore, this study evaluated the NST of CAR-T therapy using the FDA Adverse Event Reporting System (FAERS). METHODS: Data were retrieved from FAERS for the period from January 1, 2017 to March 31, 2023. Disproportionality analysis and Bayesian analysis were used for data mining. The reporting odds ratio (ROR) for NST with 95% confidence interval (CI) was calculated for each CAR-T product. The time to onset (TTO) and clinical outcomes due to CAR-T therapy-associated NST were assessed. RESULTS: Overall, 6946 cases of NST associated with CAR-T therapy were identified. The patients had a median age of 61 years (interquartile range [IQR]: 47-69 years). Significant signals were observed for all CAR-T products (ROR: 2.19, 95% CI: 2.13-2.44). Anti-CD19 CAR-T products showed a higher NST signal than anti-B cell maturation antigen (BCMA) CAR-T products (ROR025 2.13 vs. 1.98). Brexucabtagene autoleucel (ROR: 3.17, 95% CI: 2.90-3.47) and axicabtagene ciloleucel (ROR: 2.92, 95% CI: 2.81-3.03) had the two highest NST signals. For the preferred term "brain edema," the highest signals were obtained for CD28 CAR-T products. The median TTO of NST for all CAR-T products was 7 days (IQR: 3-17 days). The proportion of death, life-threatening and hospitalization adverse events associated with NST was 20.06%, 7.21%, and 32.70%, respectively. The proportion of death outcomes was higher in patients treated with tisagenlecleucel (30.36%) than in those treated with other CAR-T products, except ciltacabtagene autoleucel (P < 0.001). The proportion of hospitalizations was significantly higher for lisocabtagene maraleucel-associated NST (53.85%) than for other drugs, except for ciltacabtagene autoleucel (P < 0.001). CONCLUSIONS: NST is more closely associated with anti-CD19 CAR-Ts and CAR-Ts containing CD28. Serious NST (brain oedema) is likely to occur with CAR-Ts that contain CD28. CAR-T-related NST warrants greater attention owing to the high proportion of serious adverse events and delayed NST.