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Cancer Immunology, Immunotherapy : CII Jul 2024For advanced therapy medicinal products, the development and validation of potency assays are required, in accordance with international guidelines, to characterise the...
For advanced therapy medicinal products, the development and validation of potency assays are required, in accordance with international guidelines, to characterise the product and obtain reliable and consistent data. Our purpose was to validate the killing assay for the evaluation of autologous anti-CD19 chimeric antigen receptor (CAR) T potency. We used CD4 + and CD8 + lymphocytes or anti-CD19 CAR-T cells as effector cells and REH (CD19 +) or MOLM-13 (CD19 -) cell lines as target cells. After co-culturing target and effector cells (1:1 ratio) for 24 h, samples were labelled with 7-AAD, anti-CD3 and anti-CD19 antibodies and the frequency of CD19 + dead cells was evaluated by flow cytometry. In order to verify the CAR-T specificity for the CD19 + target, the co-culture between CAR-T and REH or MOLM-13 at different effector-to-target ratios was scheduled. Moreover, not transduced CD4 + and CD8 + lymphocytes were tested in comparison with CAR-T from the same donor to demonstrate the assay specificity. Linearity and accuracy were evaluated, and established acceptance criteria were compiled for both parameters (r ≥ 0.97 for linearity and average relative error ≤ 10% for accuracy). Furthermore, the method was considered robust when performed between 23 and 25 h of co-culture, and the intra-assay, inter-assay and inter-day precision was obtained. Finally, in order to verify the inter-analyst precision, the test was executed by three different operators and the intra-class correlation coefficient was > 0.4 in both cases. In conclusion, we consider this CAR-T potency assay as validated and usable in all steps of product development and quality control.
Topics: Humans; Receptors, Chimeric Antigen; Immunotherapy, Adoptive; Antigens, CD19; Coculture Techniques; CD8-Positive T-Lymphocytes; Cytotoxicity, Immunologic; Cell Line, Tumor; CD4-Positive T-Lymphocytes
PubMed: 38953939
DOI: 10.1007/s00262-024-03753-y -
European Journal of Nuclear Medicine... Jul 2024There is an unmet need for compounds to detect fibrillar forms of alpha-synuclein (αSyn) and 4-repeat tau, which are critical in many neurodegenerative diseases. Here,...
PURPOSE
There is an unmet need for compounds to detect fibrillar forms of alpha-synuclein (αSyn) and 4-repeat tau, which are critical in many neurodegenerative diseases. Here, we aim to develop an efficient surface plasmon resonance (SPR)-based assay to facilitate the characterization of small molecules that can bind these fibrils.
METHODS
SPR measurements were conducted to characterize the binding properties of fluorescent ligands/compounds toward recombinant amyloid-beta (Aβ), K18-tau, full-length 2N4R-tau and αSyn fibrils. In silico modeling was performed to examine the binding pockets of ligands on αSyn fibrils. Immunofluorescence staining of postmortem brain tissue slices from Parkinson's disease patients and mouse models was performed with fluorescence ligands and specific antibodies.
RESULTS
We optimized the protocol for the immobilization of Aβ, K18-tau, full-length 2N4R-tau and αSyn fibrils in a controlled aggregation state on SPR-sensor chips and for assessing their binding to ligands. The SPR results from the analysis of binding kinetics suggested the presence of at least two binding sites for all fibrils, including luminescent conjugated oligothiophenes, benzothiazole derivatives, nonfluorescent methylene blue and lansoprazole. In silico modeling studies for αSyn (6H6B) revealed four binding sites with a preference for one site on the surface. Immunofluorescence staining validated the detection of pS129-αSyn positivity in the brains of Parkinson's disease patients and αSyn preformed-fibril injected mice, 6E10-positive Aβ in arcAβ mice, and AT-8/AT-100-positivity in pR5 mice.
CONCLUSION
SPR measurements of small molecules binding to Aβ, K18/full-length 2N4R-tau and αSyn fibrils suggested the existence of multiple binding sites. This approach may provide efficient characterization of compounds for neurodegenerative disease-relevant proteinopathies.
PubMed: 38953933
DOI: 10.1007/s00259-024-06806-7 -
Intensive Care Medicine Jul 2024After cardiac surgery, fluid bolus therapy (FBT) with 20% human albumin may facilitate less fluid and vasopressor administration than FBT with crystalloids. We aimed to...
PURPOSE
After cardiac surgery, fluid bolus therapy (FBT) with 20% human albumin may facilitate less fluid and vasopressor administration than FBT with crystalloids. We aimed to determine whether, after cardiac surgery, FBT with 20% albumin reduces the duration of vasopressor therapy compared with crystalloid FBT.
METHODS
We conducted a multicentre, parallel-group, open-label, randomised clinical trial in six intensive care units (ICUs) involving cardiac surgery patients deemed to require FBT. We randomised 240 patients to receive up to 400 mL of 20% albumin/day as FBT, followed by 4% albumin for any subsequent FBT on that day, or to crystalloid FBT for at least the first 1000 mL, with use of crystalloid or 4% albumin FBT thereafter. The primary outcome was the cumulative duration of vasopressor therapy. Secondary outcomes included fluid balance.
RESULTS
Of 480 randomised patients, 466 provided consent and contributed to the primary outcome (mean age 65 years; median EuroSCORE II 1.4). The cumulative median duration of vasopressor therapy was 7 (interquartile range [IQR] 0-19.6) hours with 20% albumin and 10.8 (IQR 0-22.8) hours with crystalloids (difference - 3.8 h, 95% confidence interval [CI] - 8 to 0.4; P = 0.08). Day one fluid balance was less with 20% albumin FBT (mean difference - 701 mL, 95% CI - 872 to - 530).
CONCLUSIONS
In patients after cardiac surgery, when compared to a crystalloid-based FBT, 20% albumin FBT was associated with a reduced positive fluid balance but did not significantly reduce the duration of vasopressor therapy.
PubMed: 38953926
DOI: 10.1007/s00134-024-07488-3 -
Cancer Epidemiology, Biomarkers &... Jul 2024African Americans have higher incidence and mortality from lung cancer than non-Hispanic Whites, but investigations into differences in immune response have been...
BACKGROUND
African Americans have higher incidence and mortality from lung cancer than non-Hispanic Whites, but investigations into differences in immune response have been minimal. Therefore, we compared components of the tumor microenvironment among African Americans and non-Hispanic Whites diagnosed with non-small cell lung cancer (NSCLC) based on PD-L1 or tertiary lymphoid structure (TLS) status to identify differences of translational relevance.
METHODS
Using a cohort of 280 NSCLC patients from the INHALE study (non-Hispanic White: n=155; African American: n=125), we evaluated PD-L1 tumor proportion score (<1% vs. ≥1%) and TLS status (presence/absence), comparing differences within the tumor microenvironment based on immune cell distribution and differential expression of genes.
RESULTS
Tumors from African Americans had a higher proportion of plasma cell signatures within the tumor microenvironment than non-Hispanic Whites. In addition, gene expression patterns in African American PD-L1 positive samples suggest these tumors contained greater numbers of γδ T-cells and resting dendritic cells, along with fewer CD8+ T-cells after adjusting for age, sex, pack-years, stage, and histology. Investigation of differential expression of B-cell/plasma cell related genes between the two patient populations revealed that two immunoglobulin genes (IGKV2-29 and IGLL5) were associated with decreased mortality risk in African Americans.
CONCLUSIONS
In the first known race-stratified analysis of tumor microenvironment components in lung cancer based on PD-L1 expression or TLS status, differences within the immune cell composition and transcriptomic signature were identified that may have therapeutic implications.
IMPACT
Future investigation of racial variation within the tumor microenvironment may help direct the use of immunotherapy.
PubMed: 38953893
DOI: 10.1158/1055-9965.EPI-24-0333 -
International Journal of Systematic and... Jul 2024Two novel strain pairs (HM61/HM23 and S-34/S-58) were isolated from soil and the faeces of Tibetan antelope () collected at the Qinghai-Tibet Plateau of PR China. All...
Two novel strain pairs (HM61/HM23 and S-34/S-58) were isolated from soil and the faeces of Tibetan antelope () collected at the Qinghai-Tibet Plateau of PR China. All four new isolates were aerobic, non-motile, Gram-stain-positive, catalase-positive, oxidase-negative, and short rod-shaped bacteria. The results of phylogenetic analysis based on the full-length 16S rRNA genes and 283 core genomic genes indicated that the four strains were separated into two independent branches belonging to the genus . Strains HM61 and HM23 were most closely related to THG T63 (98.58 and 98.65 % 16S rRNA gene sequence similarity). Strains S-34 and S-58 were most closely related to MMS20-HV4-12 (98.89 and 98.89 % 16S rRNA gene sequence similarity). The G+C contents of the genomic DNA of strains HM61 and S-34 were 70.6 and 72.5 mol%, respectively. Strains HM61, S-34 and the type strains of closely related species in the analysis had average nucleotide identity values of 75.4-90.5 % as well as digital DNA-DNA hybridization values between 20.1 and 40.8 %, which clearly indicated that the four isolates represent two novel species within the genus . The chemotaxonomic characteristics of strains HM61 and S-34 were consistent with the genus . The major fatty acids of all four strains were -C, C 8 or C 9. For strains HM61 and S-34, MK-8(H) was the predominant respiratory quinone, ll-2,6-diaminopimelic acid was the diagnostic diamino acid in the cell-wall peptidoglycan, and the polar lipids profiles were composed of diphosphatidylglycerol and phosphatidylglycerol. Based on phylogenetic, phenotypic, and chemotaxonomic data, we propose that strains HM61 and S-34 represent two novel species of the genus , respectively, with the names sp. nov. and sp. nov. The type strains are HM61 (=GDMCC 4.343=JCM 36399) and S-34 (=CGMCC 4.7664=JCM 33792).
Topics: Phylogeny; RNA, Ribosomal, 16S; Soil Microbiology; Base Composition; Tibet; Fatty Acids; DNA, Bacterial; Bacterial Typing Techniques; Sequence Analysis, DNA; Feces; Antelopes; Animals; Nucleic Acid Hybridization; China; Actinomycetales; Peptidoglycan; Phospholipids
PubMed: 38953888
DOI: 10.1099/ijsem.0.006437 -
Dalton Transactions (Cambridge, England... Jul 2024Monoanionic gold bis(dithiolene) complexes were recently shown to display activity against ovarian cancer cells, Gram-positive bacteria, strains and the rodent malaria...
Monoanionic gold bis(dithiolene) complexes were recently shown to display activity against ovarian cancer cells, Gram-positive bacteria, strains and the rodent malaria parasite, . To date, only monoanionic gold(III) bis(dithiolene) complexes with a thiazoline backbone substituted with small alkyl chains have been evaluated for biomedical applications. We now analyzed the influence of the length and the hydrophobicity hydrophilicity of these complexes' alkyl chain on their anticancer and antiplasmodial properties. Isomer analogues of these monoanionic gold(III) bis(dithiolene) complexes, this time with a thiazole backbone, were also investigated in order to assess the influence of the nature of the heterocyclic ligand on their overall chemical and biological properties. In this report we present the total synthesis of four novel monoanionic gold(III) bis(dithiolene) complexes with a long alkyl chain and a polyoxygenated (PEG) chain aiming to improve their solubility and biological properties. Our results showed that the complexes with a PEG chain showed promising anticancer and antiplasmodial activities beside improved solubility, a key parameter in drug discovery and development.
PubMed: 38953883
DOI: 10.1039/d4dt01458h -
Journal of Vision Jul 2024Bayesian adaptive methods for sensory threshold determination were conceived originally to track a single threshold. When applied to the testing of vision, they do not... (Comparative Study)
Comparative Study
Bayesian adaptive methods for sensory threshold determination were conceived originally to track a single threshold. When applied to the testing of vision, they do not exploit the spatial patterns that underlie thresholds at different locations in the visual field. Exploiting these patterns has been recognized as key to further improving visual field test efficiency. We present a new approach (TORONTO) that outperforms other existing methods in terms of speed and accuracy. TORONTO generalizes the QUEST/ZEST algorithm to estimate simultaneously multiple thresholds. After each trial, without waiting for a fully determined threshold, the trial-oriented approach updates not only the location currently tested but also all other locations based on patterns in a reference data set. Since the availability of reference data can be limited, techniques are developed to overcome this limitation. TORONTO was evaluated using computer-simulated visual field tests: In the reliable condition (false positive [FP] = false negative [FN] = 3%), the median termination and root mean square error (RMSE) of TORONTO was 153 trials and 2.0 dB, twice as fast with equal accuracy as ZEST. In the FP = FN = 15% condition, TORONTO terminated in 151 trials and was 2.2 times faster than ZEST with better RMSE (2.6 vs. 3.7 dB). In the FP = FN = 30% condition, TORONTO achieved 4.2 dB RMSE in 148 trials, while all other techniques had > 6.5 dB RMSE and terminated much slower. In conclusion, TORONTO is a fast and accurate algorithm for determining multiple thresholds under a wide range of reliability and subject conditions.
Topics: Humans; Algorithms; Psychometrics; Sensory Thresholds; Visual Field Tests; Visual Fields; Bayes Theorem; Computer Simulation; Reproducibility of Results
PubMed: 38953860
DOI: 10.1167/jov.24.7.2 -
Multidisciplinary Respiratory Medicine Jul 2024Lung ultrasound (LUS) is a non-invasive point of care diagnostic tool used to assess the presence and severity of various lung disorders for more than two decades....
BACKGROUND
Lung ultrasound (LUS) is a non-invasive point of care diagnostic tool used to assess the presence and severity of various lung disorders for more than two decades. Within the healthcare professionals are the respiratory therapists (RTs) who play a vital role in managing ventilated and other patients requiring respiratory support, but the incorporation of LUS into their scope of practice has not been well highlighted. This international cross-sectional survey was specifically designed to evaluate the knowledge, attitude, and practice of RTs with respect to LUS.
METHODS
This observational cross-sectional study was conducted among RTs from different parts of the world using a questionnaire-based study tool. 514 RTs responded to all the questions and were considered for statistical analysis. Descriptive statistics, Analysis of variance, Fisher's exact, Chi-square, Bonferroni post-hoc analysis, and Binomial logistic regression analyses were performed to identify the significance of the data.
RESULTS
From 22 countries, 514 RTs responded to the survey, with the major share from the middle eastern countries. Out of the 514 responders, 44.9% of the responders were in the age group of 23-30 years; 67.1% were bachelor's degree holders; and 40.9% of participants had more than 10 years of experience. The knowledge-based questions revealed that RTs with higher experience and academic qualification provided more positive responses. While in the attitude-related domain it is observed that standardized training in LUS helps them to enhance the current practice and to add LUS to the academic curriculum of respiratory therapy schools; however there remains barriers to practice LUS based on their responses. The practice-based questions revealed that RTs expect some additional seminars/workshops/webinars to be done on LUS frequently. More than half of the participants are found to be knowledgeable with a positive attitude and working towards the inclusion of LUS in respiratory therapy profession.
CONCLUSION
It is concluded that respiratory therapists have a positive attribute towards the inclusion of lung ultrasound in their clinical practice. Providing more structured training for professional respiratory therapists and including lung ultrasound modules in the respiratory therapy school curriculum may facilitate mastering their diagnostic skills, thereby expanding the scope of practice.
PubMed: 38953856
DOI: 10.5826/mrm.2024.980 -
Nanomedicine (London, England) Jul 2024HER2, a tyrosine kinase receptor, is amplified in HER2-positive breast cancer, driving cell signaling and growth. This study aimed to combat multidrug resistance in...
HER2, a tyrosine kinase receptor, is amplified in HER2-positive breast cancer, driving cell signaling and growth. This study aimed to combat multidrug resistance in Dox-insensitive breast adenocarcinoma by creating a nanoformulation therapy with a tyrosine kinase inhibitor. Human serum albumin (HSA) was conjugated with α-D-tocopherol succinate to form nanoaggregates loaded with lapatinib (Lapa). The resulting Lapa@HSA(VE) NPs were 117.2 nm in size and demonstrated IC50 values of 10.25 μg/ml on MCF7 (S) and 8.02 μg/ml on MCF7 (R) cell lines. Lapa@HSA(VE) NPs showed no hepatotoxicity, unlike free Lapa, as seen in acute toxicity studies in rats.
PubMed: 38953854
DOI: 10.1080/17435889.2024.2359357 -
Ear and Hearing Jul 2024Evidence continues to emerge of associations between cochlear implant (CI) outcomes and cognitive functions in postlingually deafened adults. While there are multiple...
OBJECTIVES
Evidence continues to emerge of associations between cochlear implant (CI) outcomes and cognitive functions in postlingually deafened adults. While there are multiple factors that appear to affect these associations, the impact of speech recognition background testing conditions (i.e., in quiet versus noise) has not been systematically explored. The two aims of this study were to (1) identify associations between speech recognition following cochlear implantation and performance on cognitive tasks, and to (2) investigate the impact of speech testing in quiet versus noise on these associations. Ultimately, we want to understand the conditions that impact this complex relationship between CI outcomes and cognition.
DESIGN
A scoping review following Preferred Reporting Items for Systematic Reviews and Meta-Analyses guidelines was performed on published literature evaluating the relation between outcomes of cochlear implantation and cognition. The current review evaluates 39 papers that reported associations between over 30 cognitive assessments and speech recognition tests in adult patients with CIs. Six cognitive domains were evaluated: Global Cognition, Inhibition-Concentration, Memory and Learning, Controlled Fluency, Verbal Fluency, and Visuospatial Organization. Meta-analysis was conducted on three cognitive assessments among 12 studies to evaluate relations with speech recognition outcomes. Subgroup analyses were performed to identify whether speech recognition testing in quiet versus in background noise impacted its association with cognitive performance.
RESULTS
Significant associations between cognition and speech recognition in a background of quiet or noise were found in 69% of studies. Tests of Global Cognition and Inhibition-Concentration skills resulted in the highest overall frequency of significant associations with speech recognition (45% and 57%, respectively). Despite the modest proportion of significant associations reported, pooling effect sizes across samples through meta-analysis revealed a moderate positive correlation between tests of Global Cognition (r = +0.37, p < 0.01) as well as Verbal Fluency (r = +0.44, p < 0.01) and postoperative speech recognition skills. Tests of Memory and Learning are most frequently utilized in the setting of CI (in 26 of 39 included studies), yet meta-analysis revealed nonsignificant associations with speech recognition performance in a background of quiet (r = +0.30, p = 0.18), and noise (r = -0.06, p = 0.78).
CONCLUSIONS
Background conditions of speech recognition testing may influence the relation between speech recognition outcomes and cognition. The magnitude of this effect of testing conditions on this relationship appears to vary depending on the cognitive construct being assessed. Overall, Global Cognition and Inhibition-Concentration skills are potentially useful in explaining speech recognition skills following cochlear implantation. Future work should continue to evaluate these relations to appropriately unify cognitive testing opportunities in the setting of cochlear implantation.
PubMed: 38953851
DOI: 10.1097/AUD.0000000000001527