Journal: bioRxiv

Article Title: PI3K-AKT activation determines oncogenic RAS-induced hypertranscription and replication stress

doi: 10.64898/2026.03.16.711577

Figure Lengend Snippet: (A) Experimental setup for steady-state RNA sequencing. (B) Numbers of up- or downregulated genes (log2-fold change >1) after 24 or 48 h HRAS G12V or KRAS G12V induction. (C) Numbers of unique and shared up-regulated genes after HRAS G12V or KRAS G12V induction. (D) Functional enrichment analysis (gene ontology, biological process) of genes downregulated 48 h after HRAS G12V or KRAS G12V induction. (E) Functional enrichment analysis (gene ontology, biological process) of genes upregulated 48 h after HRAS G12V or KRAS G12V induction. (G) Log2 fold-change in hallmark E2F target gene expression after HRAS G12V or KRAS G12V induction. (H) E2F1 expression (RNAseq, DEseq2) after HRAS G12V or KRAS G12V induction. N=3. (I) Log2 fold-change in hallmark MYC target gene expression after HRAS G12V or KRAS G12V induction. (J) MYC expression (RNAseq, DEseq2) after HRAS G12V or KRAS G12V induction. N=3. (K) Protein levels of pRB1 and α-TUBULIN after oncogene induction for the times indicated. (L) S phase percentage after HRAS G12V induction as determined by EdU labelling and flow cytometry. N=4. (M) S phase percentage after KRAS G12V induction. N=4. (N) S phase percentage after BRAF V600E induction. N=4. (O) Experimental setup for CDK4/6 inhibitor (CDK4/6i) treatment and release. (P) S phase percentage after HRAS G12V induction and treatment with CDK4/6i. N=2. (Q) Nuclear EU intensity after HRAS G12V induction and release from CDK4/6i. N=3. (R) Average replication fork speeds after HRAS G12V induction and release from CDK4/6i. N=3. Means +/-SEM (bars) are shown with 2-way ANOVA or mixed effects analysis.

Article Snippet: Small molecule inhibitors and activators were sourced as follows: MEK inhibitor PD0325901 (1 μM) and CDK4/6 inhibitor Palbociclib (1 μM) were obtained from Merck Life Science UK Limited.

Techniques: RNA Sequencing, Functional Assay, Targeted Gene Expression, Expressing, RNA sequencing, Flow Cytometry

Journal: Molecular Oncology

Article Title: Dual PI3K / AKT and CDK4 /6 inhibition reveals selective sensitivity in an SHH medulloblastoma stem cell model

doi: 10.1002/1878-0261.70230

Figure Lengend Snippet: Impact of single and combined treatments with FGFR, PI3K, CDK4/6, and AKT inhibitors JNJ‐42756493, BYL719, PD‐0332991, and AZD5363, respectively, on the viability of NES and tNES monolayers. Cell viability of 2D NES (A–G) and tNES (H–N) cultures was assessed by absorbance measurements at 24, 48, and 72 h following treatment. Data represent independent biological replicates as follows: panel A, E, N, n = 3; panel B, D, F, (J), K, L, n = 5; panels C, n = 7; panels G, (I), n = 4; panels H, (M), n = 6. Error bars indicate standard deviation (SD). NES, neuroepithelial stem cells; tNES, tumor‐derived neuroepithelial stem cells; FGFR, fibroblast growth factor receptor; PI3K, phosphoinositide 3‐kinase; CDK4/6, cyclin‐dependent kinases 4 and 6; AKT, protein kinase B; JNJ, JNJ‐42756493; BYL, BYL719; PD, PD‐0332991; AZD, AZD5363.

Article Snippet: The FDA‐approved PI3K inhibitor BYL719 (alpelisib), AKT inhibitor AZD5363 (Capivasertib), CDK4/6 inhibitor PD‐0332991 (palbociclib), and FGFR inhibitor JNJ‐42756493 (erdafitinib) were all purchased from TargetMol (San Diego, California, USA).

Techniques: Standard Deviation, Derivative Assay, Cell Viability Assay

Journal: Molecular Oncology

Article Title: Dual PI3K / AKT and CDK4 /6 inhibition reveals selective sensitivity in an SHH medulloblastoma stem cell model

doi: 10.1002/1878-0261.70230

Figure Lengend Snippet: Impact of single and combined treatments with FGFR, PI3K, CDK4/6, and AKT inhibitors JNJ‐42756493, BYL719, PD‐0332991, and AZD5363, respectively, on the viability of NES and tNES spheroids. Cell viability of NES (A–G) and tNES (H–N) spheroids was assessed by luminescence measurements at 24, 48, and 72 h following treatment. Data represent independent biological replicates as follows: panels A, B, C, E, L, N, n = 4; panels D, H, J, M, n = 5; panels F, G, n = 3; panels I, K, n = 6. Error bars indicate standard deviation (SD). NES, neuroepithelial stem cells; tNES, tumor‐derived neuroepithelial stem cells; FGFR, fibroblast growth factor receptor; PI3K, phosphoinositide 3‐kinase; CDK4/6, cyclin‐dependent kinases 4 and 6; AKT, protein kinase B; JNJ, JNJ‐42756493; BYL, BYL719; PD, PD‐0332991; AZD, AZD5363.

Article Snippet: The FDA‐approved PI3K inhibitor BYL719 (alpelisib), AKT inhibitor AZD5363 (Capivasertib), CDK4/6 inhibitor PD‐0332991 (palbociclib), and FGFR inhibitor JNJ‐42756493 (erdafitinib) were all purchased from TargetMol (San Diego, California, USA).

Techniques: Spheroids, Standard Deviation, Derivative Assay, Cell Viability Assay

Journal: Molecular Oncology

Article Title: Dual PI3K / AKT and CDK4 /6 inhibition reveals selective sensitivity in an SHH medulloblastoma stem cell model

doi: 10.1002/1878-0261.70230

Figure Lengend Snippet: Evaluation of drug interactions between BYL719, PD‐0332991, and AZD5363 in NES and tNES spheroids 72 h after treatment. Drug interaction analyses were performed in NES and tNES spheroids using the SynergyFinder Plus platform. Heatmap plots (A–D) and 3D surface maps (E–H) display the interaction profiles and optimal combination doses of BYL719, PD‐0332991, and AZD5363 based on the Highest Single Agent (HSA) model. The synergy score reflects the interaction: values greater than 10 indicate a synergistic effect, values between −10 and 10 suggest an additive effect, and values below −10 indicate an antagonistic effect. NES, neuroepithelial stem cells; tNES, tumor‐derived neuroepithelial stem cells; BYL, BYL719; PD, PD‐0332991; AZD, AZD5363.

Article Snippet: The FDA‐approved PI3K inhibitor BYL719 (alpelisib), AKT inhibitor AZD5363 (Capivasertib), CDK4/6 inhibitor PD‐0332991 (palbociclib), and FGFR inhibitor JNJ‐42756493 (erdafitinib) were all purchased from TargetMol (San Diego, California, USA).

Techniques: Spheroids, Derivative Assay

Journal: Molecular Oncology

Article Title: Dual PI3K / AKT and CDK4 /6 inhibition reveals selective sensitivity in an SHH medulloblastoma stem cell model

doi: 10.1002/1878-0261.70230

Figure Lengend Snippet: The efficacy of the FGFR, PI3K, CDK4/6, and AKT inhibitors JNJ‐42756493, BYL719, PD‐0332991, and AZD5363, respectively, alone and in combination, on the growth of NES and tNES spheroids. Growth of 3D NES (A–G) and 3D tNES (H–N) was monitored every 2 h for 72 h following treatment. Values were normalized to the PBS control within each experiment and treatment condition. Data represent independent biological replicates as follows: panels A, B, G, H, and I, n = 3; panels C, D, E, and F, n = 5; panels J, K, L, and M, n = 6; and panel N, n = 4. Error bars indicate the standard deviation of independent biological replicates. NES, neuroepithelial stem cells; tNES, tumor‐derived neuroepithelial stem cells; FGFR, fibroblast growth factor receptor; PI3K, phosphoinositide 3‐kinase; CDK4/6, cyclin‐dependent kinases 4 and 6; AKT, protein kinase B; JNJ, JNJ‐42756493; BYL, BYL719; PD, PD‐0332991; AZD, AZD5363.

Article Snippet: The FDA‐approved PI3K inhibitor BYL719 (alpelisib), AKT inhibitor AZD5363 (Capivasertib), CDK4/6 inhibitor PD‐0332991 (palbociclib), and FGFR inhibitor JNJ‐42756493 (erdafitinib) were all purchased from TargetMol (San Diego, California, USA).

Techniques: Spheroids, Control, Standard Deviation, Derivative Assay

Journal: Molecular Oncology

Article Title: Dual PI3K / AKT and CDK4 /6 inhibition reveals selective sensitivity in an SHH medulloblastoma stem cell model

doi: 10.1002/1878-0261.70230

Figure Lengend Snippet: The efficacy of the FGFR, PI3K, CDK4/6, and AKT inhibitors JNJ‐42756493, BYL719, PD‐0332991, and AZD5363, respectively, alone and in combination, on the growth of NES and tNES spheroids. Representative images of 3D NES (A1–30) and 3D tNES (B1–30) are shown 72 h after treatment from three independent biological experiments ( n = 3). Scalebar = 400 μm. NES, neuroepithelial stem cells; tNES, tumor‐derived neuroepithelial stem cells; FGFR, fibroblast growth factor receptor; PI3K, phosphoinositide 3‐kinase; CDK4/6, cyclin‐dependent kinases 4 and 6; AKT, protein kinase B; JNJ, JNJ‐42756493; BYL, BYL719; PD, PD‐0332991; AZD, AZD5363.

Article Snippet: The FDA‐approved PI3K inhibitor BYL719 (alpelisib), AKT inhibitor AZD5363 (Capivasertib), CDK4/6 inhibitor PD‐0332991 (palbociclib), and FGFR inhibitor JNJ‐42756493 (erdafitinib) were all purchased from TargetMol (San Diego, California, USA).

Techniques: Spheroids, Derivative Assay

Journal: Molecular Oncology

Article Title: Dual PI3K / AKT and CDK4 /6 inhibition reveals selective sensitivity in an SHH medulloblastoma stem cell model

doi: 10.1002/1878-0261.70230

Figure Lengend Snippet: Histological analysis of NES and tNES spheroids 48 h after treatment with single doses of BYL719, PD‐0332991, AZD5363 and their combinations. Representative images of NES and tNES spheroid sections stained for the proliferation marker Ki67 (A) and the apoptosis marker cleaved caspase‐3 (CC‐3) (B) are shown. Ki67 staining is shown for NES (A1–4 and A9–11) and tNES (A5–8 and A12–14), and CC‐3 staining is shown for NES (B1–4 and B9–11) and tNES (B5–8 and B12–14). Images were captured at 20x magnification. Experiments were repeated independently three times ( n = 3), with similar results. Scalebar = 100 μm. NES, neuroepithelial stem cells; tNES, tumor‐derived neuroepithelial stem cells; BYL, BYL719; PD, PD‐0332991; AZD, AZD5363.

Article Snippet: The FDA‐approved PI3K inhibitor BYL719 (alpelisib), AKT inhibitor AZD5363 (Capivasertib), CDK4/6 inhibitor PD‐0332991 (palbociclib), and FGFR inhibitor JNJ‐42756493 (erdafitinib) were all purchased from TargetMol (San Diego, California, USA).

Techniques: Analysis, Spheroids, Staining, Marker, Derivative Assay, Immunohistochemistry