Seven Supplementary Figures and Legends

Chromatin structure in enzalutamide treated C4-2 cells

Supplementary Figure 3 from Ligand-Dependent Platelet-Derived Growth Factor Receptor (PDGFR)-α Activation Sensitizes Rare Lung Cancer and Sarcoma Cells to PDGFR Kinase Inhibitors

AbstractAbiraterone is a standard treatment for metastatic castrate-resistant prostate cancer (mCRPC) that slows disease progression by abrogating androgen synthesis and antagonizing the androgen receptor (AR). Here we report that inhibitors of the mitotic regulator polo-like kinase-1 (Plk1), including the clinically active third-generation Plk1 inhibitor onvansertib, synergizes with abiraterone in vitro and in vivo to kill a subset of cancer cells from a wide variety of tumor types in an androgen-independent manner. Gene-expression analysis identified an AR-independent synergy-specific gene set signature upregulated upon abiraterone treatment that is dominated by pathways related to mitosis and the mitotic spindle. Abiraterone treatment alone caused defects in mitotic spindle orientation, failure of complete chromosome condensation, and improper cell division independently of its effects on AR signaling. These effects, although mild following abiraterone monotherapy, resulted in profound sensitization to the antimitotic effects of Plk1 inhibition, leading to spindle assembly checkpoint-dependent mitotic cancer cell death and entosis. In a murine patient-derived xenograft model of abiraterone-resistant metastatic castration-resistant prostate cancer (mCRPC), combined onvansertib and abiraterone resulted in enhanced mitotic arrest and dramatic inhibition of tumor cell growth compared with either agent alone. Overall, this work establishes a mechanistic basis for the phase II clinical trial (NCT03414034) testing combined onvansertib and abiraterone in mCRPC patients and indicates this combination may have broad utility for cancer treatment.Significance:Abiraterone treatment induces mitotic defects that sensitize cancer cells to Plk1 inhibition, revealing an AR-independent mechanism for this synergistic combination that is applicable to a variety of cancer types.

Supplementary methods. Supplementary Table S1. Verification of the analytical sensitivity (lower limit of detection) of the KRASG12/G13 mutation-enrichment NGS assay. Supplementary Table S2. KRASG12/G13 mutations in archival tumor tissue, urine cell-free DNA (cfDNA) and plasma cfDNA. Supplementary Table S3. Systemic therapies in patients with serial urine and/or plasma cell-free DNA (cfDNA) collection. Supplementary Figure S1. Workflow and characteristics of the platform used to analyze cell-free DNA in urine and plasma. Supplementary Figure S2. Schematic of the mutation-enrichment next-generatin sequencing (NGS) assay for the detection of KRASG12/13 mutations in cell-free DNA (cfDNA). Supplementary Figure S2. Schematic of the mutation-enrichment next-generatin sequencing (NGS) assay for the detection of KRASG12/13 mutations in cell-free DNA (cfDNA). Supplementary Figure S3. Kaplan-Meier curves of overall survival (OS) based on the number of KRASG12/13 copies in cell-free DNA (cfDNA). Supplementary Figure S4. Kaplan-Meier curves of overall survival (OS) based on the concentration of cell-free DNA (cfDNA).

Supplementary Figure 4 from Ligand-Dependent Platelet-Derived Growth Factor Receptor (PDGFR)-α Activation Sensitizes Rare Lung Cancer and Sarcoma Cells to PDGFR Kinase Inhibitors

Supplemental Table 1: Concordance of initial urinary cell-free DNA (cfDNA) assessment of BRAFV600E mutation with tissue biopsy BRAFV600E result.

Supplemental Figure 1: BRAFV600E mutant allele burden in cell-free DNA (cfDNA) of urine and plasma from treatment naïve patients based on BRAFV600E tissue genotype result.

Supplementary Figures 1-7 - PDF file 117K, Supplementary Figure 1. Cohorts used for BCI model training and validation Supplementary Figure 2. Performance of H:I and MGI for early (0-5 years) and late (> 5 years) distant recurrence in Stockholm ER+, LN- untreated patients. A, C: H:I, 0-5 years and > 5 years. B, D: MGI, 0-5 years and > 5 years. Supplementary Figure 3. BCI risk groups for prediction of overall distant recurrence in Stockholm TAM and Multi-institutional cohorts, ER+, LN- patients. Supplementary Figure 4. Continuous BCI score for overall (0 - 10 years) rate of distant recurrence in Stockholm TAM cohort. Supplementary Figure 5. BCI risk groups for prediction of early and late distant recurrence in the combined StockholmTAM and Multi-institutional cohort; ER+, LN-, tamoxifen-treated patients. Supplementary Figure 6. Continuous BCI score for early (0 - 5 years) and late (> 5 years) rate of distant recurrence in the combined Stockholm TAM and Multi-institutional cohort (ER+, LN-, tamoxifen-treated patients). Supplementary Figure 7. Prediction of tamoxifen benefit by H/I. A: no benefit of tamoxifen in H/I-low patients. B: benefit of tamoxifen in H/I-high patients

Supplementary Figures 1-2 from Ligand-Dependent Platelet-Derived Growth Factor Receptor (PDGFR)-α Activation Sensitizes Rare Lung Cancer and Sarcoma Cells to PDGFR Kinase Inhibitors

Supplementary Data from A Five-Gene Molecular Grade Index and HOXB13:IL17BR Are Complementary Prognostic Factors in Early Stage Breast Cancer

Supplementary Methods, Figures 1-8, Tables 1-3 from A Genome-Wide Screen for Microdeletions Reveals Disruption of Polarity Complex Genes in Diverse Human Cancers

Supplementary Figure 5 from Ligand-Dependent Platelet-Derived Growth Factor Receptor (PDGFR)-α Activation Sensitizes Rare Lung Cancer and Sarcoma Cells to PDGFR Kinase Inhibitors

Supplemental Figure 2: Radiographic, histologic, and molecular evaluation of KRASG12S mutant patient with Erdheim-Chester Disease (ECD).

Supplementary Figure 7 from Ligand-Dependent Platelet-Derived Growth Factor Receptor (PDGFR)-α Activation Sensitizes Rare Lung Cancer and Sarcoma Cells to PDGFR Kinase Inhibitors

Supplemental Table 2: Concordance of initial plasma cell-free DNA (cfDNA) assessment of BRAFV600E mutations with tissue biopsy BRAFV600E result.

A: Analysis of dilution series of indicated mutant EGFR variants spiked into 60 ng (â‰^18,180 genome equivalents) of WT DNA. Each data point represents one preparative within 6 independent dilutions series prepared and analyzed by two operators on two different instruments on three non-consecutive days for a total of 18 samples per dilution point. An analysis algorithm was applied to transform the mutant EGFR sequencing reads into the absolute mutant copies detected. The box-and-whisker plots show the median (center line), 25th and 75th percentiles (box) with the connecting "whiskers" extending from the first quartile minus 1.5 of the interquartile range (IQR, the third quartile less the first quartile) and the third quartile plus 1.5 of the IQR. A positive Spearman's correlation close to 1 indicates a strong, positive relationship between the absolute mutant EGFR copies detected and the absolute mutant EGFR copies per input. B: Inter-run reproducibility of the EGFR exon 19 deletions, L858R and T790M enrichment PCR-NGS assays for the dilution series shown in panel A. The Coefficient of Variation Percent (CV%) was calculated as the ratio of the standard deviation to the mean of the absolute EGFR copies detected within each absolute copy per input level and is reported as a percentage.

Detailed list of variants identified by targeted-NGS

Urine volumes, DNA concentrations, DNA input amounts, the number of output mutant sequencing reads, mutant copies detected and the 95% Confidence Intervals

Supplementary Tables and Figure