human kinome crispr knockout library preparation  (Addgene inc)

Journal: bioRxiv

Article Title: Polypharmacology of an Optimal Kinase Library

doi: 10.64898/2026.03.17.711623

Figure Lengend Snippet: (a) The number of OKL inhibitors ( KS Kd < 100 nM) per kinase shown on the kinome tree by the size and intensity of the markers. Kinases not inhibited are outlined in red, kinases not assayed are outlined in black. (b) Radial bar plot showing the number of OKL inhibitors with KS Kd ≤ 100 nM for all mutant kinases (red bars) assayed in the KINOMEscan panel. WT kinases are indicated with blue bars. (c) The maximum partition index (PI max ) for each inhibitor in OKL. The inset barplots show the most (top) and least (bottom) selective inhibitors and associated kinase target colored by current clinical status. (d) PI max for each kinase. The inset CORAL tree shows the PI max for each kinase, the darker and redder the circle, the more selectively that kinase is inhibited by an OKL compound. (e) Boxplot showing the number of kinases per inhibitor with KS Kd ≤ 100 nM by clinical status. (f) Boxplot showing the maximum partition index for each OKL inhibitor by clinical status. P-value is from a Welch’s t-test comparing tool compounds to all others. (g) Histogram of the KS Kd values for the assigned target(s) of OKL compounds. (h) Histogram of the partition indices for the assigned target(s) of OKL compounds. (i) Cumulative stairstep plot showing how often the nominal target is in the top N targets by KS Kd for approved (green) and not approved (gray) inhibitors. (j) Scatterplot showing PI max with respect to the year each compound was first published. The lines of best fit to all data points and to the highest datapoint per year are shown in solid red and dashed maroon, respectively. (k) Histogram showing the number of OKL inhibitors that bind each dark and illuminated kinase with KS Kd ≤ 100 nM. Boxplots representing the same data are shown below the histograms. The * indicates Welch’s t-test P < 0.0001.

Article Snippet: The KINOMEscan® kinome-scale profiling assay available from Eurofins is 5-10-fold cheaper and has therefore been used in more than 2,000 publications.

Techniques: Mutagenesis

Journal: bioRxiv

Article Title: Polypharmacology of an Optimal Kinase Library

doi: 10.64898/2026.03.17.711623

Figure Lengend Snippet: (a) The number of inhibitors with KS Kd ≤ 1 µM per kinase. The inset plot shows the top 20 kinases most widely inhibited by OKL compounds colored by group. (b) The number of OKL inhibitors ( KS Kd < 1 µM) per kinase shown on the kinome tree by the size and intensity of the markers. (c) Radial bar plot showing the number of OKL inhibitors with KS Kd ≤ 1 µM for all mutant kinases (red bars) assayed in the KINOMEscan panel. WT kinases are indicated with blue bars. (d) Boxplot showing the number of kinases per inhibitor with KS Kd ≤ 100 nM by maximum stage of clinical development reached. (e) Boxplot showing the maximum partition index for each OKL inhibitor by maximum stage of clinical development reached. P-value is from a one-way ANOVA with Tukey’s correction for multiple comparisons. (f) Histogram of the maximum partition indices for all dark and light kinases. KDE lines are overlaid for visualization only.

Article Snippet: The KINOMEscan® kinome-scale profiling assay available from Eurofins is 5-10-fold cheaper and has therefore been used in more than 2,000 publications.

Techniques: Mutagenesis

Journal: bioRxiv

Article Title: Polypharmacology of an Optimal Kinase Library

doi: 10.64898/2026.03.17.711623

Figure Lengend Snippet: (a) Look-up table based on screening goal and assay concentration showing the sensitivity (SENS), specificity (SPEC) and precision (PREC) for the optimal cutoff values. (b) Kinome tree showing the classification of each kinase in a KINOMEscan of 10 µM ruxolitinib using a percent control threshold of 22. (c) Kinome tree showing the kinases that are misclassified using a 35% threshold and correctly classified using a 22% threshold. (d) Hexagonally binned scatter plots showing the KS Kd values estimated from all two dose pairs with respect to those estimated from all four doses. (e) Summary barplot of the mean squared error (MSE) of log 10 (K d ) for all compound–kinase pairs for KS Kd values estimated from all two dose pairs.

Article Snippet: The KINOMEscan® kinome-scale profiling assay available from Eurofins is 5-10-fold cheaper and has therefore been used in more than 2,000 publications.

Techniques: Concentration Assay, Control

Journal: NPJ Breast Cancer

Article Title: CDK2 inhibition enhances CDK4/6 inhibitor antitumor activity in comprehensive breast cancer PDX model screen

doi: 10.1038/s41523-025-00851-7

Figure Lengend Snippet: a BLU-222 and palbociclib proliferative GI 50 determined by CyQUANT direct proliferation assay, 5-day compound treatment. BLU-222 responder threshold was set at 200 nM as previously described . Dashed lines connect the BLU-222 and palbociclib IC 50 s from an individual cell line. P -value determined by paired t -test. b Volcano plot assessing differential gene expression in BLU-222 responders versus palbociclib responders from cell lines in ( a ). Points above the horizontal dashed line have an adjusted P < 0.05. The left vertical dashed line and the right vertical dashed line represent log 2 (FC) = -1.5 and log 2 (FC) = 1.5, respectively. c Kinome-selectivity of CDK2 inhibitors, as measured by KINOMEscan at 3 μM compound. S(10) is the number of kinases inhibited at <10% of control, divided by the total number of human wild-type kinases. Kinome illustrations reproduced courtesy of Cell Signaling Technology, Inc. ( www.cellsignal.com ) (CSTI). The foregoing website is maintained by CSTI, and Blueprint Medicines is not responsible for its content. d Antiproliferative response to CDK2 inhibitors in breast cancer cell lines as in ( a ). Cell lines were categorized by RB1 (Rb) expression (+: intact, −: <25th percentile expression with copy number loss or mutation), CDKN2A (p16) expression (+: >75th percentile of mRNA expression, -: ≤25th percentile of mRNA expression), and CCNE1 (cyclin E1) expression (overexpression defined as above the 75th percentile of mRNA expression). CCNE1 copy number status is noted by symbol color. Statistical significance was evaluated using the Wilcoxon rank sum test. e–l CyQuant, 5-day compound treatment, in T-47D isogenic cell lines overexpressing CCNE1 , CDKN2A , or both. Mean and standard error of the mean (SEM) of two biological replicates are plotted. m Cell cycle distribution in T-47D isogenic cell lines in response to 24-h treatment BLU-222 (100 nM) or palbociclib (250 nM), ±siRB1. Mean and standard deviation (SD) of two biological replicates are plotted. n Antitumor activity of BLU-222 (60 mg/kg twice a day [BID]), ribociclib (50 mg/kg once a day [QD]), and BLU-222+ribociclib (60 mg/kg BID + 50 mg/kg QD) in BCX.022, a TNBC PDX model. 2-way ANOVA statistical deviation in tumor volume at study end is noted, * P < 0.05, ** P < 0.01. Mean tumor volume and SEM are plotted, n = 6 per group.

Article Snippet: Kinome illustrations reproduced courtesy of Cell Signaling Technology, Inc. ( www.cellsignal.com ) (CSTI).

Techniques: CyQUANT Assay, Proliferation Assay, Gene Expression, Control, Expressing, Mutagenesis, Over Expression, Standard Deviation, Activity Assay