Journal: Cell Reports Medicine

Article Title: CDK4/6 inhibition overcomes venetoclax resistance mechanisms with enhanced combination activity in acute myeloid leukemia

doi: 10.1016/j.xcrm.2025.102526

Figure Lengend Snippet: Venetoclax, in combination with the CDK4/6 inhibitor palbociclib, inhibits growth, stalls cell cycle progression, and reduces tumor burden in AML models (A–C) OCI-AML2 cells assessed after a 5-day treatment with palbociclib (1 μM), venetoclax (200 nM), or the combination (equimolar to single agents) to determine levels of apoptosis (A), cell cycle progression (B), and percentage of viable OCI-AML2 cells/mL of media (C) Data represent the mean ± SD for 3 replicates (∗∗ p ≤ 0.01). (D) Schematic depicting two independent PDX experiments. PDX model 1: evaluation of disease burden after injection of AML patient tumor cells. PDX model 2: survival studies after injection of AML patient tumor cells. The schematic was generated using BioRender. (E–G) Flow cytometry analysis at time of euthanization for PDX model 1 showing human (h)CD45 blasts in peripheral blood (E), spleen weight of animals (F), and percentage of hCD45 chimerism in spleen tissue (G). Mean values ± SEM are shown unless otherwise stated. Two-tailed Student’s t tests were used for comparisons (∗ p ≤ 0.05, ∗∗ p ≤ 0.01, ∗∗∗ p ≤ 0.001, and ∗∗∗∗ p ≤ 0.0001). (H) Survival Kaplan-Meier curves for PDX model 2 (log rank [Mantel-Cox] test, ∗ p ≤ 0.05, ∗∗ p ≤ 0.01, ∗∗∗ p ≤ 0.001, and ∗∗∗∗ p ≤ 0.0001).

Article Snippet: Human AML cell lines (OCI-AML2, OCI-AML3, MOLM13) were purchased from DSMZ.

Techniques: Injection, Generated, Flow Cytometry, Two Tailed Test

Journal: Cell Reports Medicine

Article Title: CDK4/6 inhibition overcomes venetoclax resistance mechanisms with enhanced combination activity in acute myeloid leukemia

doi: 10.1016/j.xcrm.2025.102526

Figure Lengend Snippet: A combination of ven+palbo leads to changes in protein synthesis rate and translational machinery (A) Bulk RNA-seq for 560 primary AML samples, showing Pearson correlations between AML cellular state eigengenes (columns) and Reactome Translation pathway eigengenes (rows). Protein synthesis pathways positively correlate (red) with a progenitor-like cell-state signature and negatively correlate (blue) with a monocyte-like cell-state signature. (B) MTS assays confirming drug responsiveness in parental/venetoclax-sensitive cells (VenS/Par) and venetoclax-resistant cells (VenR) for OCI-AML2 cell lines. Data points denote the mean normalized cell viability ± SD for 3 replicates. (C) SUnSET assay to assess protein synthesis in OCI-AML2 cells treated with drug for 24 h. Puromycin incorporation into newly synthesized proteins was measured in ven-sensitive (VenS/Par [parental]) and -resistant (VenR) cells. A representative image of n = 4 immunoblots is shown. (D) Estimation plots of intensity measurements of anti-puromycin signal from 4 separate immunoblot experiments. (E) Immunoblots of OCI-AML2 cells following 24-h or 5-day drug treatments. Vinculin is used as a loading control. (F) SUnSET assay immunoblots showing puromycin incorporation in progenitor-like or monocyte-like patient samples after drug treatment. (G) Flow-cytometry-based SUnSET assay detecting CD64 + , CD11b + , CD33 + monocytes. The percentage of puromycin-positive cells reflects active protein synthesis within this monocyte population.

Article Snippet: Human AML cell lines (OCI-AML2, OCI-AML3, MOLM13) were purchased from DSMZ.

Techniques: RNA Sequencing, Synthesized, Western Blot, Control, Flow Cytometry

Journal: Cell Reports Medicine

Article Title: CDK4/6 inhibition overcomes venetoclax resistance mechanisms with enhanced combination activity in acute myeloid leukemia

doi: 10.1016/j.xcrm.2025.102526

Figure Lengend Snippet: Loss of IKZF1 leads to increased expression of AXL and is increased in IKZF1 -mutated AML patient samples (A) Volcano plot highlighting genes of interest from RNA-seq in IKZF1 -KO OCI-AML2 cells, color coded based on primary known function. (B) qPCR validation of RNA-seq results showing mean fold change ± SD for 3 replicates. (C) Immunoblot shows upregulation of AXL protein with loss of IKZF1 in OCI-AML2 cells. (D) AXL mRNA is overexpressed in AML primary patient samples harboring IKZF1 mutations ( n = 9) compared to WT samples ( n = 662). (E) OCI-AML2 IKZF1 -KO cells show resistance (red dots) to palbo, ven, and ven+palbo and retain drug sensitivity to several AXL inhibitors (blue dots). Sensitivity is shown as a percentage of the maximum area under the dose response curve (AUC) derived for a 7-point concentration series ranging from 10 μM to 10 nM. (F) AUC values from ex vivo drug sensitivity assays for 4 primary AML samples, each harboring the IKZF1 hotspot mutation N159S (∗ p ≤ 0.05, ∗∗ p ≤ 0.01, ∗∗∗ p ≤ 0.001, and ∗∗∗∗ p ≤ 0.0001 by Student’s t test).

Article Snippet: Human AML cell lines (OCI-AML2, OCI-AML3, MOLM13) were purchased from DSMZ.

Techniques: Expressing, RNA Sequencing, Biomarker Discovery, Western Blot, Derivative Assay, Concentration Assay, Ex Vivo, Mutagenesis