Abstract 6731: Oncogenic ASPP2k: a key regulator of classical hallmarks of cancer in triple negative breast cancer (TNBC)

Abstract ASPP2 is a tumor suppressor, originally characterized as direct p53-binding protein. More recently, ASPP2 was shown to also directly interact with BCL-2 and NFkB, putting ASPP2 in a central position to orchestrate major cancer related pathways. In this context, we have recently reported the identification of a novel, oncogenic isoform of ASPP2, named ASPP2kappa (k), which is highly and frequently expressed in TNBC. As ASPP2k is characterized by loss of the p53- as well as partial loss of BCL-2 and NFkB-binding sites, its expression impairs major pathways controlling cellular fate, resulting in accelerated proliferation and resistance to chemotherapy. We now show that expression of ASPP2k modulates all classical hallmarks of cancer - fueling a highly aggressive tumor biology in TNBC. Importantly we demonstrate that silencing of ASPP2k slows down tumor growth and inhibits metastasis in vivo. Employing our isogenic TNBC ASPP2κ interferenced (KD) cell models (MDA-MB-231 and HS-578T, n=18 per model) we studied the role of ASPP2κ in NSG xenotransplant models. Mice were xenotransplanted with EV or ASPP2k KD cells and tumor engraftment, progression, angiogenesis and metastasis were monitored using an IVIS Lumina imaging system. Tumor engraftment and growth were found to be significantly attenuated in the KD strains (representative data for MDA model: doubling times EV: 7.3d; KD: 12.8d; p<0, 0001). EV mice had to be sacrificed between day 40-60 after injection due to tumor size (median OS 51d) while ASPP2k KD only presented with small, localized tumors at day of termination (d72, OS<0, 0001). Importantly, 100% of mice bearing ASPP2k expressing (EV) tumors, presented with distant metastasis in multiple different sites at time of sacrifice, while in ASPP2k KD mice, metastasis could be detected in max. one site and only 30% of mice. Regulation of pro-angiogenic and EMT markers as well as involvement of the NFkB pathway were confirmed to play a role in ASPP2k-mediated metastasis. Most importantly, we were able to silence ASPP2k in vivo, employing novel, fully chemically modified ASPP2k-specific siRNAs generated for in vivo use and confirm potent inhibition of tumor growth also in these models. In summary, we here provide absolutely novel insights into TNBC tumor biology in dependency of ASPP2k and demonstrate first successful attempts to silence ASPP2κ employing highly specific siRNAs to target ASPP2k in vivo. Citation Format: Alessia Ruiba, Anna-Lena Ahrens, Lara Aldinger, Lea Stiehl, Reka Haraszti, Wolfram Jochum, Christoph Driessen, Marcus M. Schittenhelm, Kerstin Maria Kampa-Schittenhelm. Oncogenic ASPP2k: a key regulator of classical hallmarks of cancer in triple negative breast cancer (TNBC) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 6731.