OC44 - Therapeutic potential of adavosertib (AZD1775), a WEE1 kinase inhibitor, in advanced adrenocortical carcinoma: focus on molecular changes

Background/Introduction Adrenocortical carcinoma (ACC) is a rare endocrine tumor, often diagnosed at advanced stages and associated with poor prognosis. Treatment options for advanced ACC are limited, highlighting the need for novel therapeutic strategies. The WEE1 inhibitor adavosertib (AZD1775) could offer a novel approach, as it promotes mitotic progression in the presence of DNA damage, leading to mitotic catastrophe in cancer cells. Purpose Here, we evaluate the WEE1 expression in adrenal tissues and the effects of adavosertib on in vitro ACC models as monotherapy and in combination with currently used DNA-damaging agents. Methods WEE1 mRNA expression was analyzed by qRT-PCR in 91 adrenal tissues (19 normal adrenal glands [NAG], 20 adenomas [ACA], 52 ACC). WEE1 immunohistochemistry was performed in 114 ACC samples and correlated with clinical outcomes. Expression at RNA and protein levels was further assessed in five ACC cell lines. Cytotoxicity of adavosertib, alone and combined with cisplatin or gemcitabine, was evaluated in vitro using CellTiter-Glo and Western blot analysis. Results WEE1 mRNA levels were significantly higher in ACC compared to NAG (0.032 vs 0.012, p=0.04) but not to ACA. High WEE1 protein staining was associated with shorter progression free survival (HR=2.67, 95%CI=0.74-9.66, p=0.07) and overall survival (HR=2.11, 95%CI=0.48-9.37, p=0.32) at Cox-regression analysis together with ENSAT tumor stage. WEE1 expression was found in all ACC cell lines, with CU-ACC1 having the lowest expression at both mRNA and protein levels. Adavosertib induced cytotoxicity in NCI-H295R, JIL-2266, and CU-ACC2 cells (IC50: 1.17, 1.35, and 0.4 µM, respectively), while CU-ACC1 and MUC-1 were less responsive. Combination with cisplatin or gemcitabine showed an additive-synergic effect across all cell lines. At molecular level, adavosertib reduced CDK1 phosphorylation and increased markers of DNA damage (pH2AX), stress response (pRPA32), and apoptosis (cleaved PARP) after 24h and 48h of treatment compared to control (DMSO treatment). Conclusion(s) Our findings demonstrate elevated WEE1 expression in ACC, suggesting its promising role as therapeutic target as well as potential prognostic marker. Adavosertib exhibits cytotoxic activity in ACC models, enhanced by combination with DNA-damaging agents. We will further investigate WEE1 inhibition by including proteomic approaches to elucidate resistance and sensitivity mechanisms.