Development of optimized cytotoxicity assays for assessing the antitumor potential of CAR-T cells.

CAR-T cells are T cells expressing a chimeric antigen receptor (CAR) rendering them capable of killing tumor cells after recognition of a target antigen. CD19 CAR-T cells have revolutionized the treatment of hematological malignancies. Their function is typically assessed by cytotoxicity assays using human allogeneic cell lines expressing the target antigen CD19 such as Nalm-6. However, an alloreactive reaction is observed with these cells, leading to a CD19-independent killing. To address this issue, we developed a fluorescence microscopy-based potency assay using murine target cells to provide an optimized cytotoxicity assay with enhanced specificity towards CD19. Murine NIH/3T3 (3T3) fibroblast-derived cell line and EL4 T-cell lymphoma-derived cell line were used as targets (no xenoreactivity was observed after coculture with human T cells). 3T3 and EL4 cells were engineered to express eGFP (enhanced Green Fluorescent Protein) and CD19 or CD22 using retroviral vectors. CD19 CAR-T cells and non-transduced (NT) control T cells were produced from several donors. After 4 h or 24 h, alloreactive cytotoxicity against CD19+ Nalm-6-GFP cells and CD19- Jurkat-GFP cells was observed with NT or CAR-T cells. In the same conditions, CAR-T but not NT cells specifically killed CD19+ but not CD19- 3T3-GFP or EL4-GFP cells. Both microscope- and flow cytometry-based assays revealed as sensitive as impedance-based assay. Using flow cytometry, we could further determine that CAR-T cells had mostly a stem cell-like memory phenotype after contact with EL4 target cells. Therefore, CD19+ 3T3-GFP or EL4-GFP cells and fluorescence microscopy- or flow cytometry-based assays provide convenient, sensitive and specific tools to evaluate CAR-T cell function with no alloreactivity.

A comparison of COVID-19 incidence rates across six European countries in 2021.

International comparisons of COVID-19 incidence rates have helped gain insights into the characteristics of the disease, benchmark disease impact, shape public health measures and inform potential travel restrictions and border control measures. However, these comparisons may be biased by differences in COVID-19 surveillance systems and approaches to reporting in each country. To better understand these differences and their impact on incidence comparisons, we collected data on surveillance systems from six European countries: Belgium, England, France, Italy, Romania and Sweden. Data collected included: target testing populations, access to testing, case definitions, data entry and management and statistical approaches to incidence calculation. Average testing, incidence and contextual data were also collected. Data represented the surveillance systems as they were in mid-May 2021. Overall, important differences between surveillance systems were detected. Results showed wide variations in testing rates, access to free testing and the types of tests recorded in national databases, which may substantially limit incidence comparability. By systematically including testing information when comparing incidence rates, these comparisons may be greatly improved. New indicators incorporating testing or existing indicators such as death or hospitalisation will be important to improving international comparisons.

Correction: Andrographis paniculata (Chuan Xin Lián) for symptomatic relief of acute respiratory tract infections in adults and children: A systematic review and meta-analysis.

[This corrects the article DOI: 10.1371/journal.pone.0181780.].

Andrographis paniculata (Chuan Xin Lián) for symptomatic relief of acute respiratory tract infections in adults and children: A systematic review and meta-analysis.

INTRODUCTION: Antimicrobial resistance (AMR) is a substantial threat to public health. Safe and effective alternatives are required to reduce unnecessary antibiotic prescribing. Andrographis Paniculata (A. Paniculata, Chuan Xin Lián) has traditionally been used in Indian and Chinese herbal medicine for cough, cold and influenza, suggesting a role in respiratory tract infections (RTIs). This systematic review aimed to evaluate the clinical effectiveness and safety of A. Paniculata for symptoms of acute RTIs (ARTIs). MATERIALS AND METHODS: English and Chinese databases were searched from their inception to March 2016 for randomised controlled trials (RCTs) evaluating oral A. Paniculata without language barriers (Protocol ID: CRD42016035679). The primary outcomes were improvement in ARTI symptoms and adverse events (AEs). A random effects model was used to pool the mean differences and risk ratio with 95% CI reported. Methodological quality was evaluated using the Cochrane risk of bias tool; two reviewers independently screened eligibility and extracted data. RESULTS: Thirty-three RCTs (7175 patients) were included. Most trials evaluated A. Paniculata (as a monotherapy and as a herbal mixture) provided commercially but seldom reported manufacturing or quality control details. A. Paniculata improved cough (n = 596, standardised mean difference SMD: -0.39, 95% confidence interval CI [-0.67, -0.10]) and sore throat (n = 314, SMD: -1.13, 95% CI [-1.37, -0.89]) when compared with placebo. A. Paniculata (alone or plus usual care) has a statistically significant effect in improving overall symptoms of ARTIs when compared to placebo, usual care, and other herbal therapies. Evidence also suggested that A. Paniculata (alone or plus usual care) shortened the duration of cough, sore throat and sick leave/time to resolution when compared versus usual care. No major AEs were reported and minor AEs were mainly gastrointestinal. The methodological quality of included trials was overall poor. CONCLUSIONS: A. Paniculata appears beneficial and safe for relieving ARTI symptoms and shortening time to symptom resolution. However, these findings should be interpreted cautiously owing to poor study quality and heterogeneity. Well-designed trials evaluating the effectiveness and potential to reduce antibiotic use of A. Paniculata are warranted.