Figure 4.
Figure 4. Newer-generation HSP90 inhibitors potently inhibit MCL in vitro and in vivo. (A) Cell-cycle analysis by flow cytometry upon treatment with AUY922 50 nM after 0, 8, and 24 hours, and Annexin V (AV) and PI staining of MCL lines treated as indicated. Data are representative of 2 to 4 independent experiments. (B) ROS as measured by DCFDA fluorescence intensity at 3 hours for Mino, Jeko-1, and MAVER-1 cell lines cultured with DCFDA along with vehicle, TBHP (positive control), or AUY922. Error bars indicate SD. ***P < .001, ****P < .0001. (C) IC50 values for 10 primary MCL samples. (D) Mice were inoculated subcutaneously with either MAVER-1 or Z-138 (n = 40 per line) and treated with AUY922 or vehicle once a volume of 100 to 200mm3 was reached. Error bars indicate standard error of the mean (SEM). P = .001 for both MAVER-1 and Z-138 compared with vehicle by 2-way ANOVA. (E) 18F-FLT PET imaging of mice with Z-138 xenografts treated for 5 days. SUVtotal indicates the standard uptake value integrated over the entire tumor volume. (F) Images from 18F-FLT PET in NSG mice xenografted with primary patient MCL and treated with either vehicle or AUY922 for 5 days. (G) Average SUVmax (n = 3 per arm) for the indicated organs. Proximal femur and lymph nodes each represent averages of right and left within the same animal. P values based on 2-sided Student t test. Error bars indicate SD. (H) Distribution of proteins that were upregulated (red) or downregulated (green) by log2 fold ≥0.5 following treatment with AUY922 in mouse PDX models and 4 of 5 or 5 of 5 cell lines. (I) Violin plots of protein fold changes for proteins that were altered by log2 fold ≥0.5 in the mouse PDX model alone and in both the mouse PDX model and 1 to 5 cell lines following treatment with AUY922 vs DMSO. (J) Immunoblotting of lysates from spleens of mice xenografted with primary MCL and treated for 5 days, and log fold change of each of protein represented in the immunoblot from the proteomic experiment. ALN, axillary lymph node; B, bladder; GIT, gastrointestinal tract; K, kidney; PF, proximal femur; S, spleen.

Newer-generation HSP90 inhibitors potently inhibit MCL in vitro and in vivo. (A) Cell-cycle analysis by flow cytometry upon treatment with AUY922 50 nM after 0, 8, and 24 hours, and Annexin V (AV) and PI staining of MCL lines treated as indicated. Data are representative of 2 to 4 independent experiments. (B) ROS as measured by DCFDA fluorescence intensity at 3 hours for Mino, Jeko-1, and MAVER-1 cell lines cultured with DCFDA along with vehicle, TBHP (positive control), or AUY922. Error bars indicate SD. ***P < .001, ****P < .0001. (C) IC50 values for 10 primary MCL samples. (D) Mice were inoculated subcutaneously with either MAVER-1 or Z-138 (n = 40 per line) and treated with AUY922 or vehicle once a volume of 100 to 200mm3 was reached. Error bars indicate standard error of the mean (SEM). P = .001 for both MAVER-1 and Z-138 compared with vehicle by 2-way ANOVA. (E) 18F-FLT PET imaging of mice with Z-138 xenografts treated for 5 days. SUVtotal indicates the standard uptake value integrated over the entire tumor volume. (F) Images from 18F-FLT PET in NSG mice xenografted with primary patient MCL and treated with either vehicle or AUY922 for 5 days. (G) Average SUVmax (n = 3 per arm) for the indicated organs. Proximal femur and lymph nodes each represent averages of right and left within the same animal. P values based on 2-sided Student t test. Error bars indicate SD. (H) Distribution of proteins that were upregulated (red) or downregulated (green) by log2 fold ≥0.5 following treatment with AUY922 in mouse PDX models and 4 of 5 or 5 of 5 cell lines. (I) Violin plots of protein fold changes for proteins that were altered by log2 fold ≥0.5 in the mouse PDX model alone and in both the mouse PDX model and 1 to 5 cell lines following treatment with AUY922 vs DMSO. (J) Immunoblotting of lysates from spleens of mice xenografted with primary MCL and treated for 5 days, and log fold change of each of protein represented in the immunoblot from the proteomic experiment. ALN, axillary lymph node; B, bladder; GIT, gastrointestinal tract; K, kidney; PF, proximal femur; S, spleen.

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