Journal: Blood Advances

Article Title: CAR T cells targeting CCR4 selectively deplete human Tregs ex vivo and in vivo

doi: 10.1182/bloodadvances.2025017573

Figure Lengend Snippet: Tregs are depleted by the CCR4-CAR in a humanized mouse model. (A) Experimental design. NSG-SGM3-IL15 engrafted with CD34 + hematopoietic stem cells were injected with 1 million CAR + CCR4-CARTs IV. Blood was collected on days 0, 3, 5, and 8, and mice were euthanized on day 11. (B) Representative flow plots showing the proportion of human CD45 (hCD45) and mCD45 leukocytes at baseline. (C) Proportion of Tregs, CD4 + non-Treg, and CD4 − cells of hCD45 percent at baseline. (D) Percentage of Tregs, non-Treg, and CD4 − cells that are CCR4 + at baseline. (E) Representative flow plots showing the CCR4 + and FOXP3 + expression on the CD4 + population before and after CART administration gated on CD4 + cells. (F-K) Proportions of Tregs, CD4 + non-Tregs, and CD4 − cells over time. Significance was determined using t tests corrected for multiple comparisons, with comparison to baseline indicated on graph; ∗ P < .05; ∗∗ P < .01; ∗∗∗ P < .001; ∗∗∗∗ P < .0001. M1, Mouse 1; M2, Mouse 2; M3, Mouse 3; mCD45, mouse CD45.

Article Snippet: Mice were engrafted with human cord blood–derived CD34 + hematopoietic stem cells, and females were available for use at age 13 to 18 weeks after evaluation of engrafted human cell populations by flow cytometry at The Jackson Laboratory.

Techniques: Injection, Expressing, Comparison

Journal: eBioMedicine

Article Title: Effectorless Fc-fusion improves FLT3L drug-like properties for cancer immunotherapy combinations

doi: 10.1016/j.ebiom.2025.105822

Figure Lengend Snippet: In vitro characterisation of FLT3L-Fc NG2LH activity . Representative sensorgrams of FLT3L-Fc NG2LH binding to recombinant human monomeric FLT3-monoFc (a) and dimeric FLT3-Fc (b). Black solid lines are curves fit to the data using a 1:1 binding model, and coloured lines are raw data. The tested recombinant human FLT3-monoFc concentrations (from bottom to top) are 62.5, 125, 250, 500, 1000, and 2000 nM; recombinant human dimeric FLT3-Fc concentrations (from bottom to top) are 3.75, 7.5, 15, 30, 60, and 120 nM. The sensorgrams were generated after in-line reference cell correction followed by buffer sample subtraction. The experiments were conducted at 37 °C. (c) Binding affinities of recombinant human monomeric FLT3-monoFc and dimeric FLT3-Fc proteins to captured FLT3L-Fc NG2LH. k a association rate constant; k d dissociation rate constant; K D equilibrium dissociation constant. Data for FLT3L-Fc NG2LH binding to recombinant human FLT3-monoFc were averaged from three independent experimental Biacore runs. Data for FLT3L-Fc NG2LH binding to recombinant human dimeric FLT3-Fc were averaged from two independent experimental Biacore runs. (d) ADCP for FLT3L-Fc NG2LH and its variants. Data shown are a representative set taken from one of three independent experiments. Average (SD) of duplicate values are shown. (e) Proliferation of OCI-AML5 cells in vitro. A representative dose response curve is shown, Average (SD) of triplicate values. Data were normalised to 10 μg/mL FLT3L-Fc NG2LH as the maximum (100%) response and the assay media alone control as the minimum (0%) response. (f) Human cDC1 differentiation from cord blood stem cells. Frequency of cDC1 at the end of culture is shown (percent of live cells). Results were obtained from 2 donors. (g) In vitro proliferation of mouse bone marrow cells in response to mFLT3L or mFLT3L-Fc. Average (SD) of duplicate values are shown.

Article Snippet: CD34+ human cord blood stem cells (StemCell Technologies) were seeded at 5000 cells per well in 96-well U-bottom plates (Costar) in 100 μL expansion media (StemSpan media–StemCell Technologies, 10% FBS, 40 ng/mL IL-3, 200 ng/mL SCF, 100 ng/mL TPO–cytokines from Peprotech).

Techniques: In Vitro, Activity Assay, Binding Assay, Recombinant, Generated, Control