Fig. 7: BMAL1-HIF2α heterodimers are sensitive to disruption by PT2399.

A Detection of HA-tagged stabilized HIF2α and FLAG-tagged class I bHLH-PAS proteins by immunoblot of lysates (input) or complexes purified with an anti-FLAG antibody from HEK 293 cells expressing the indicated plasmids and treated with the indicated concentrations of PT2399 for 1 h. B Quantitation of data like that shown in (A), normalized to 0 μM control groups. Data represent the mean ± s.d. for n = 3 biological replicates. C Relative luminescence units detected in HEK293T cells expressing luciferase under the control of a hypoxia-responsive element with overexpressed stabilized HIF2α and ARNT (salmon) or BMAL1 (green) and treated with the indicated concentrations of PT2399. Data show mean ± s.d. for n = 5 biological replicates from one of three experiments with similar results. **** P < 1e⁻⁶. D, E Detection of BMAL1, NR1D1, SLC2A14, and DDIT4 transcripts in patient-derived xenograft tumors collected from mice treated with PT2399 (red) or vehicle control (black) grouped by whether tumor growth was reduced by PT2399 (sensitive) or not (resistant) (D) or in 786O cells expressing the indicated shRNAs and treated with 10 μM PT2399 (red) or vehicle control (black) for 6 h (E). Data represent transcripts per million detected by sequencing RNA collected from 11 resistant or 12 sensitive PDX tumors (D) or the mean ± s.e.m. for relative expression normalized to RPLP0 by quantitative RT-PCR for three biological replicates each measured in triplicate (E). *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001 by two-way ANOVA or mixed effects analysis with Tukey’s correction for multiple hypothesis testing. Source data are provided as a Source Data file.