Journal: Cancer research

Article Title: CDK2 Inhibition Exerts RB-Independent Antitumor Activity in CDK4/6 Inhibitor–Resistant HR + /HER2 − Breast Cancer

doi: 10.1158/0008-5472.CAN-25-1859

Figure Lengend Snippet: CDK2 inhibition does not target RB in most of the HR + /HER2 − breast cancers. A and B, Western blot analysis with the indicated antibodies of lysates from the patient-derived HR + MBC cell lines MGH10035.1, MGH358.1, and MGH406.4, and the CCNE1 -amplified TNBC MDA-MB-157 and ovarian cancer OVCAR3 cell lines treated with different concentrations of CDK4/6 (palbociclib), CDK2/4/6 (PF-06873600), or CDK2 (PF-07104091) inhibitors. C, In vitro kinase assay using recombinant cyclin A2/CDK1, cyclin A2/CDK2, and cyclin D3/CDK2 after incubation with increased concentrations of PF-07104091 CDK2i. Data were normalized to incubation with DMSO, and error bars indicate standard deviations ( n = 3). IC 50 values were 1.370 nmol/L for cyclin A2/CDK1, 0.0821 nmol/L for cyclin A2/CDK2, and 3.062 nmol/L for cyclin D3/CDK2. IC 50 (nmol/L) was calculated using nonlinear regression of the dose–response inhibition. D and E, Cell viability of MGH10035.1 ( C ) and MCF7 ( D ) cells expressing shSCR or shRB1 and treated with CDK4/6is, CDK2/4/6is, or CDK2is for 7 days. Data were normalized to DMSO-treated cells. Error bars indicate standard deviations ( n = 3). F, Heatmaps presenting the CDK4/6i, CDK2/4/6i, or CDK2i log IC 50 (nmol/L) in MGH10035.1, MCF7, CAMA1, MDA-MB-361, and MDA-MB-415 HR + breast cancer cell lines. IC 50 was calculated using nonlinear regression of the dose–response inhibition.

Article Snippet: CDK2 (cat. #sc-163, RRID: AB_631215), cyclin D1 (cat. #sc-753, RRID: AB_2070433), and cyclin E1 (cat. #sc-247, RRID: AB_627357) antibodies were purchased from Santa Cruz Biotechnology, and the tubulin (cat. #T9026, RRID: AB_477593) antibody was purchased from Sigma-Aldrich.

Techniques: Inhibition, Western Blot, Analysis, Derivative Assay, Amplification, In Vitro, Kinase Assay, Recombinant, Incubation, Expressing

Journal: Oncogenesis

Article Title: Cyclin E modulates vulnerability to CDC7 kinase inhibition

doi: 10.1038/s41389-026-00613-5

Figure Lengend Snippet: A Representative western blot of HCC-1806 cells for cyclin E, CDK2, and pCDK1 (Y15) following CCNE1 or CDK2 knockdown. B Representative cell cycle profiling of HCC-1806 cells following CCNE1 knockdown and 48 hr treatment with indicated concentrations of TAK-931. C Quantification of cell cycle state from HCC-1806 cells following CCNE1 knockdown and 48 hr treatment with indicated concentrations of TAK-931 (n = 3; two-way ANOVA with Tukey’s multiple comparisons test accounting for gene and drug effect with significance determined for ≥ 4 N DNA; error bars represent SD). D HCC-1806 cells following CCNE1 knockdown and treatment with 125 nM (left) or 250 nM (right) TAK-931 for five days and monitored with Cellcyte or Incucyte live cell proliferative software ( n = 4; two-way ANOVA with Tukey’s multiple comparisons test accounting for gene and drug effect using endpoint normalized GFP counts; error bars represent SEM). E IC 50 curves derived from live cell proliferative analyses as described in ( D ) ( n = 4; two-way ANOVA with Tukey’s multiple comparisons test accounting for gene and drug effect using endpoint normalized GFP counts; error bars represent SEM). F Representative images captured at the end of five-day live cell proliferative analyses from HCC-1806 cells following CCNE1 knockdown and treatment with 125 nM TAK-931. G Representative colony outgrowth assay from HCC-1806 cells with CCNE1 CRISPR knockout or non-targeting control and treatment with indicated concentrations of TAK-931 for three cycles, with treatment replenishment every five days (left) and western blot of HCC-1806 CRISPR CCNE1 isogenic pair for cyclin E, CDK2, and pCDK1 (Y15) (right).

Article Snippet: Antibodies were obtained from the following vendors: Ph.RB (S807/811) (Cell Signaling Technology Cat# 8516, RRID:AB_11178658), CDK2 (Cell Signaling Technology Cat# 2546, RRID:AB_2276129), Ph.CDK1 (Y15) (Cell Signaling Technology Cat# 9111, RRID:AB_331460), CDK1 (Cell Signaling Technology Cat# 77055, RRID:AB_2716331), Cyclin B1 (Cell Signaling Technology Cat# 12231, RRID:AB_2783553), Cleaved PARP (Cell Signaling Technology Cat# 9542, RRID:AB_2160739), Ph.STAT1 (S727) (Cell Signaling Technology Cat# 8826, RRID:AB_2773718), STAT1 (Cell Signaling Technology Cat# 9172, RRID:AB_2198300), Ph.CHK1 (S317) (Cell Signaling Technology Cat# 2344, RRID:AB_331488), Ph.CHK1 (S345) (Cell Signaling Technology Cat# 2348, RRID:AB_331212), CHK1 (Cell Signaling Technology Cat# 2360, RRID:AB_2080320), CDK1 (Santa Cruz Biotechnology Cat# sc-54, RRID:AB_627224) (1:500), MCM-7 (Santa Cruz Biotechnology Cat# sc-9966, RRID:AB_627235) (1:500), Ph.MCM2 (S40/41) (Abcam Cat# ab70371, RRID:AB_2141957), Cyclin E (Bethyl Cat# A301-566A, RRID:AB_1039994), MCM-4 (Thermo Fisher Scientific Cat# MA5-52516, RRID:AB_3248993), β-actin (R and D Systems Cat# MAB8929, RRID:AB_3076436).

Techniques: Western Blot, Knockdown, Software, Derivative Assay, CRISPR, Knock-Out, Control

Journal: Oncogenesis

Article Title: Cyclin E modulates vulnerability to CDC7 kinase inhibition

doi: 10.1038/s41389-026-00613-5

Figure Lengend Snippet: A Representative western blot of T-47D (left) and MDA-MB-231 (right) cells for cyclin E, CDK2, and pCDK1 (Y15) following CCNE1 induction. B Representative cell cycle profiling of T-47D cells following CCNE1 induction and 48 hr treatment with indicated concentrations of TAK-931. C Quantification of cell cycle state from T-47D cells following CCNE1 induction and 48 hr treatment with indicated concentrations of TAK-931 ( n = 3; two-way ANOVA with Tukey’s multiple comparisons test accounting for gene and drug effect with significance determined for ≥ 4 N DNA; error bars represent SD). D Representative cell cycle profiling of MDA-MB-231 cells following CCNE1 induction and 48 hr treatment with indicated concentrations of TAK-931. E Quantification of cell cycle state from MDA-MB-231 cells following CCNE1 induction and 48 hr treatment with indicated concentrations of TAK-931 ( n = 3; two-way ANOVA with Tukey’s multiple comparisons test accounting for gene and drug effect with significance determined for ≥ 4 N DNA; error bars represent SD). F T-47D cells following CCNE1 induction and treatment with 15.625 nM (left) or 31.25 nM (right) TAK-931 for five days and monitored with Cellcyte or Incucyte live cell proliferative software ( n = 4; two-way ANOVA with Tukey’s multiple comparisons test accounting for gene and drug effect using endpoint normalized GFP counts; error bars represent SEM). G IC 50 curves derived from live cell proliferative analyses as described in ( F ) ( n = 4; two-way ANOVA with Tukey’s multiple comparisons test accounting for gene and drug effect using endpoint normalized GFP counts; error bars represent SEM). H , MDA-MB-231 cells following CCNE1 induction and treatment with 62.5 nM (left) or 125 nM (right) TAK-931 for five days and monitored with Cellcyte or Incucyte live cell proliferative software ( n = 4; two-way ANOVA with Tukey’s multiple comparisons test accounting for gene and drug effect using endpoint normalized GFP counts; error bars represent SEM). I IC 50 curves derived from live cell proliferative analyses as described in ( H ) ( n = 4; two-way ANOVA with Tukey’s multiple comparisons test accounting for gene and drug effect using endpoint normalized GFP counts; error bars represent SEM). J Representative colony outgrowth assays from T-47D (left) and MDA-MB-231 (right) cells following CCNE1 induction and treatment with indicated concentrations of TAK-931 for three cycles, with doxycycline and treatment replenishment every five days. K Representative BrdU incorporation analysis of T-47D cells following CCNE1 induction and 48 hr treatment with indicated concentrations of TAK-931.

Article Snippet: Antibodies were obtained from the following vendors: Ph.RB (S807/811) (Cell Signaling Technology Cat# 8516, RRID:AB_11178658), CDK2 (Cell Signaling Technology Cat# 2546, RRID:AB_2276129), Ph.CDK1 (Y15) (Cell Signaling Technology Cat# 9111, RRID:AB_331460), CDK1 (Cell Signaling Technology Cat# 77055, RRID:AB_2716331), Cyclin B1 (Cell Signaling Technology Cat# 12231, RRID:AB_2783553), Cleaved PARP (Cell Signaling Technology Cat# 9542, RRID:AB_2160739), Ph.STAT1 (S727) (Cell Signaling Technology Cat# 8826, RRID:AB_2773718), STAT1 (Cell Signaling Technology Cat# 9172, RRID:AB_2198300), Ph.CHK1 (S317) (Cell Signaling Technology Cat# 2344, RRID:AB_331488), Ph.CHK1 (S345) (Cell Signaling Technology Cat# 2348, RRID:AB_331212), CHK1 (Cell Signaling Technology Cat# 2360, RRID:AB_2080320), CDK1 (Santa Cruz Biotechnology Cat# sc-54, RRID:AB_627224) (1:500), MCM-7 (Santa Cruz Biotechnology Cat# sc-9966, RRID:AB_627235) (1:500), Ph.MCM2 (S40/41) (Abcam Cat# ab70371, RRID:AB_2141957), Cyclin E (Bethyl Cat# A301-566A, RRID:AB_1039994), MCM-4 (Thermo Fisher Scientific Cat# MA5-52516, RRID:AB_3248993), β-actin (R and D Systems Cat# MAB8929, RRID:AB_3076436).

Techniques: Western Blot, Software, Derivative Assay, BrdU Incorporation Assay

Journal: Oncogenesis

Article Title: Cyclin E modulates vulnerability to CDC7 kinase inhibition

doi: 10.1038/s41389-026-00613-5

Figure Lengend Snippet: A Representative western blot of HCC-1806 cells for total MCM-4, 7, and cyclin E following CCNE1 knockdown. B Representative flow cytometric MCM-4 chromatin binding assay from HCC-1806 cells following CCNE1 knockdown and 48 hr treatment with 125 nM TAK-931. C Representative flow cytometric MCM-7 chromatin binding assay from HCC-1806 cells following CCNE1 knockdown and 48 hr treatment with 125 nM TAK-931. D Representative flow cytometric MCM-4 chromatin binding assay from T-47D cells following CCNE1 induction and 48 hr treatment with 62.5 nM TAK-931. E Representative flow cytometric MCM-7 chromatin binding assay from T-47D cells following CCNE1 induction and 48 hr treatment with 62.5 nM TAK-931.

Article Snippet: Antibodies were obtained from the following vendors: Ph.RB (S807/811) (Cell Signaling Technology Cat# 8516, RRID:AB_11178658), CDK2 (Cell Signaling Technology Cat# 2546, RRID:AB_2276129), Ph.CDK1 (Y15) (Cell Signaling Technology Cat# 9111, RRID:AB_331460), CDK1 (Cell Signaling Technology Cat# 77055, RRID:AB_2716331), Cyclin B1 (Cell Signaling Technology Cat# 12231, RRID:AB_2783553), Cleaved PARP (Cell Signaling Technology Cat# 9542, RRID:AB_2160739), Ph.STAT1 (S727) (Cell Signaling Technology Cat# 8826, RRID:AB_2773718), STAT1 (Cell Signaling Technology Cat# 9172, RRID:AB_2198300), Ph.CHK1 (S317) (Cell Signaling Technology Cat# 2344, RRID:AB_331488), Ph.CHK1 (S345) (Cell Signaling Technology Cat# 2348, RRID:AB_331212), CHK1 (Cell Signaling Technology Cat# 2360, RRID:AB_2080320), CDK1 (Santa Cruz Biotechnology Cat# sc-54, RRID:AB_627224) (1:500), MCM-7 (Santa Cruz Biotechnology Cat# sc-9966, RRID:AB_627235) (1:500), Ph.MCM2 (S40/41) (Abcam Cat# ab70371, RRID:AB_2141957), Cyclin E (Bethyl Cat# A301-566A, RRID:AB_1039994), MCM-4 (Thermo Fisher Scientific Cat# MA5-52516, RRID:AB_3248993), β-actin (R and D Systems Cat# MAB8929, RRID:AB_3076436).

Techniques: Western Blot, Knockdown, Binding Assay

Journal: Cell Death & Disease

Article Title: Dual targeting of PI3Kδ and PPARα enhances antitumor activity via FoxO1 activation in follicular lymphoma

doi: 10.1038/s41419-026-08593-5

Figure Lengend Snippet: A - C EdU flow plots for Karpas-422, RL, and Sc-1 cells after 24-h treatment (Linperlisib 32 μM; Chiglitazar 16 μM). D – F Quantification of EdU-positive cells showing reduced DNA synthesis with the combination ( n = 3). G – L ( G , H ) for Karpas-422; ( I , J ) for RL; ( K , L ) for Sc-1. Cell-cycle distribution by PI staining, demonstrating G0/G1 accumulation with Linperlisib and further enhancement by the combination (Linperlisib 16 μM; Chiglitazar 8 μM; n = 3). M GSEA indicating negative enrichment of the G1/S transition pathway (NES = − 2.85865, p < 0.00001). N – P Western blots of G1/S checkpoint proteins (p27, Cyclin E1, CDK2, p-CDK2); combination treatment increased p27 and decreased Cyclin E1, CDK2, and p-CDK2 (representative of three independent experiments).

Article Snippet: The primary antibodies used were as follows: PI3K p110δ (A19742; ABclonal), pan-Akt (#4691; CST), phospho-Akt (Ser473, #4060; CST), PPARα (ab227074; Abcam), FoxO1 (#2880; CST), phospho-FoxO1 (Thr24, #9464; CST) and (Ser256, #9461; CST), FoxO3a (#12829; CST), FoxO4 (#9472; CST), Mcl-1 (#16225-1-AP; Proteintech), Bcl-2 (#12789-1-AP; Proteintech), Bim (A19702; ABclonal), Bax (#50599-2-Ig; Proteintech), Cleaved PARP(#5625; CST), P21(#2947; CST), p27 (#3686; CST), Cyclin E1 (#11935-1-AP; Proteintech), CDK2 (#10122-1-AP; Proteintech), phospho-CDK2 (#4539; CST), GLUT1(#73015; CST), PGK1(A12686; ABclonal), β-Tubulin (#10094-1-AP; Proteintech), Lamin B1 (#13435; CST), and β-Actin (#66009-1-Ig; Proteintech).

Techniques: DNA Synthesis, Staining, Western Blot