Three Supplementary Tables and Supplementary Figures S1-S13 from SAR405838: An Optimized Inhibitor of MDM2–p53 Interaction That Induces Complete and Durable Tumor Regression

Abstract

<p>SI Table S1. Crystallography data collection and refinement statistics for the co-crystal structure of SAR405838 complexed with human MMD2 protein. SI Table S2. Binding affinities of SAR405838, as well as reference compounds to different MDM2 protein constructs, as determined using competitive FP-based binding assays. SI Table S3. Key pharmacokinetic parameters for SAR405838 in mice, rats and dogs with intravenous (i.v.) or oral (p.o.) administration. SI Figure S1. Binding of SAR405838 to MDMx, Bcl-2, Bcl-xL, Mcl-1 and β-catenin. SI Figure S2. Chemical stability of SAR405838 in three different solutions. SI Figure S3. qRT-PCR Analysis of mRNA levels of p53-targeted genes (MDM2, p21, PUMA, NOVA and BAX) in SJSA-1 and HCT-116 cells treated with SAR405838 (SAR), MI-219 and nutlin-3a. SI Figure S4. Western blot analysis of accumulation of p53, MDM2, PUMA, p21 proteins and cleavage of PARP and caspase-3 induced by SAR405838 (SAR) and MI-219 in prostate cancer LNCAP and acute leukemia RS4;11 cell lines. SI Figure S5. Western blot analysis of p53, MDM2, PUMA and p21 protein level treated with different concentration of SAR405838 (SAR) for 24 hrs in representative cell lines with p53 mutation or deletion. SI Figure S6. Stable knock-down of p53 by shRNAi in SJSA-1 cell line effectively attenuates cell growth inhibition and cell cycle arrest. SI Figure S7. Stable knock-down of p53 by shRNAi in RS4;11 cell line effectively attenuates cell growth inhibition and apoptosis induction by SAR405838. SI Figure S8. Stable knock-down of p53 by shRNAi in LNCaP cell line effectively attenuates cell growth inhibition and cell cycle arrest by SAR405838. SI Figure S9. Western blot analysis of p53, MDM2, p21, PUMA, PARP and caspase-3 in SJSA-1 tumors treated with a single, oral dose of SAR405838 at 100 mg/kg. SI Figure S10. (a). qRT-PCR Analysis of mRNA expression of MDM2, p21 and PUMA in HCT-116 tumors treated with SAR405838. (b). Analysis of protein levels of p53, MDM2, p21, cleaved PARP and caspase-3 in HCT-116 tumors treated with SAR405838 using a Mesoscale assay. SI Figure S11. (a). qRT-PCR Analysis of mRNA expression of MDM2, p21 and PUMA in RS4;11 tumors treated with SAR405838. (b). Analysis of protein levels of p53, MDM2, p21, cleaved PARP and caspase-3 in RS4;11 tumors treated with SAR405838 using a Mesoscale assay. SI Figure S12. (a). qRT-PCR Analysis of mRNA expression of MDM2, p21 and PUMA in LNCaP tumors treated with a single dose of SAR405838. (b). Analysis of protein levels of p53, MDM2, p21, cleaved PARP and caspase-3 in LNCaP tumors treated with a single dose of SAR405838 using a Mesoscale assay. SI Figure S13 Mouse weights in efficacy experiments in SJSA-1, HCT-116, RS4;11 and LNCaP xenograft models.</p>