Supplementary material, methods and data from Dose and Schedule Determine Distinct Molecular Mechanisms Underlying the Efficacy of the p53–MDM2 Inhibitor HDM201
Authors: Jeay Sébastien , Ferretti Stéphane , Holzer Philipp , Fuchs Jeanette , Chapeau Emilie A. , Wartmann Markus , Sterker Dario , Romanet Vincent , Murakami Masato , Kerr Grainne , Durand Eric Y. , Gaulis Swann , Cortes-Cros Marta , Ruetz Stephan , Stachyra Therese-Marie , Kallen Joerg , Furet Pascal , Würthner Jens , Guerreiro Nelson , Halilovic Ensar , Jullion Astrid , Kauffmann Audrey , Kuriakose Emil , Wiesmann Marion , Jensen Michael R. , Hofmann Francesco , Sellers William R. 
Published: April 2023

<p>Supplementary material and methods include the description for the bioanalytical method for HDM201 detection in plasma and tumor, the human and mouse gene expression analysis in vitro and in vivo, the live-cell quantification of cleaved-caspase activation, the western blot analysis, the immunohistochemistry, the splinkerette PCR for the amplification of transposon integration sites and the tumor sequencing, mapping of insertion sequences to the mouse genome and identification of common integration site, additional information on the shRNA screen and the tumor models and supplementary references. Supplementary figures include: Ã¢â‚¬Â¢	Fig S1: the SJSA-1 inhibition growth curves when treated with HDM201 at different doses and for different times and the data for MOLM-3. Ã¢â‚¬Â¢	Fig S2: the cumulative percentage of cleaved-caspase-3/7 positive cells over the time, the GI50 of HDM201, CGM097 or nutlin-3a on SJSA-1 cells and the cellular apoptosis, as judged by AUC of cleaved-caspase-3/7 positive cells, induced by these compounds. Ã¢â‚¬Â¢	Fig S3: the PK profile in plasma and tumor of HDM201 in SJSA-1 tumors-bearing rat after p.o. and i.v. treatment, the Bcl-xl mRNA levels in tumors after HDM201 treatment, representative images of SJSA-1 tumors stained with p53 and cleaved-caspase 3 antibodies after HDM201 treatment and the individual data for the efficacy experiment in SJSA-1 tumor-bearing rats. Ã¢â‚¬Â¢	Fig S4: the PD of HDM201 in PB tumor bearing nude mice after single dose administration. Supplementary tables include: Ã¢â‚¬Â¢	Table S1: Biochemical profile of HDM201. Ã¢â‚¬Â¢	Table S2: List of cell lines tested for their sensitivity to HDM201 (n=291) Ã¢â‚¬Â¢	Table S3: Contingency table indicating association between sensitivity to HDM201 and TP53 wild-type status. Ã¢â‚¬Â¢	Table S4: List of cell lines tested for their sensitivity to both MDM2 knock-down by shRNA and HDM201 (n=261) Ã¢â‚¬Â¢	Table S5: Contingency table indicating association between sensitivity to HDM201 and sensitivity to MDM2 shRNA. Ã¢â‚¬Â¢	Table S6: List of significant rescuer and sensitizer genes following both HDM201 treatment types Ã¢â‚¬Â¢	Table S7: Pharmacokinetic parameters for HDM201 after p.o. and i.v. dosing in rat. Ã¢â‚¬Â¢	Table S8: Summary of primary PK parameters for HDM201 daily regimen after single dose (Day 1) in patients. Ã¢â‚¬Â¢	Table S9: Summary of primary PK parameters for HDM201 daily regimen on Day 14 in patients. Ã¢â‚¬Â¢	Table S10: Summary of primary PK parameters for HDM201 q3w regimen after single dose in patients.</p>