Fig. 1: Combined PARP and WEE1 inhibitor treatment induces synergistic anti-tumour effects as well as STING-mediated inflammatory response in BRCA1/2 wild-type TNBC.

a Synergy quantitation via Chou-Talalay combination index (CI) values of olaparib and AZD1775 in TNBC cell lines. All cell lines are BRCA1/2 wild type except MDA-MB-436 (BRCA1 mutant indicated by *). Grey bar: CI values indicating additive effect. BL: Basal-like; MSL; Mesenchymal-like; LAR: Luminal androgen receptor. Data depicts mean ± s.d. b Images show immunofluorescence of γH2AX in MDA-MB-231 treated with DMSO (V), olaparib (O) and/or AZD1775 (A) for 72 h. Scale bar represents 50 µm. Graph shows mean fold change relative to vehicle controls ± s.d. analysed via flow cytometry. c Assessing protein expression of replication stress markers via Western blot after 24 h of indicated treatment. d Assessing protein expression of substrates in the STING pathway via Western blot after 24 (pIRF3 and pTBK1) and 72 (cGAS) hours of indicated treatment. Results in (c) and (d) are representative of at least 2 independent experiments. *: The GAPDH expression marked by * is the loading control for both pIRF3 Ser369 and pTBK1 Ser172 of the MDA-MB-231 cell line. e Gene expression via qRT-PCR in HCC1806 and MDA-MB-231 cells after 72 h treatment with olaparib and AZD1775. Data shows mean relative mRNA expression ± s.d. f Comparing gene expression levels in HCC1806 and MDA-MB-231 cells with STING knockout (sgTMEM173) vs sgAAVS1 control cells via qRT-PCR after 72 h treatment with olaparib and AZD1775. Data shows mean relative mRNA expression ± s.d. g Top ranked, upregulated GSEA Hallmarks in HCC1806 and MDA-MB-231 cells treated in vitro for 72 h with olaparib and AZD1775. Normalized P < 0.05, FDR < 0.25. h HCC1806 and MDA-MB-231 were treated for 72 h with vehicle, olaparib, AZD1775, and/or interferon (I; 5 ng/mL) and analysed via flow cytometry for tumour cell-surface expression of major histocompatibility complex (MHC) markers HLA-ABC and HLA-DR. Data depicts mean fold change of mean fluorescent intensity (MFI) relative to vehicle controls ± s.d. *P < 0.05; **P < 0.01; ***P < 0.001; ****P < 0.0001 by one-way ANOVA.