Abstract A035: Targeting ISCA2 and Fe-S assembly to induce ferroptosis in kidney cancer

Abstract Background: Clear cell renal cell carcinoma (ccRCC) is typically initiated by VHL loss, resulting in constitutive activation of HIF-1α and HIF-2α. VHL loss/HIF activation promote metabolic reprogramming that increases sensitivity to ferroptosis, a form of cell death associated with iron-dependent lipid peroxidation. However, current ferroptosis inducers are not suitable drug candidates. A screen for HIF-2α inhibitors identified compounds that target Iron Sulfur Cluster Assembly 2 (ISCA2), a protein required for the maturation of a subset of mitochondrial iron-sulfur [4Fe-4S] proteins. ISCA2 inhibition triggers the iron starvation response, facilitating iron uptake and death via ferroptosis, as well as decreased HIF-1/2α translation. We previously described compound #25 as an inhibitor of ISCA2 which decreases HIF-1/2α levels and induces ferroptosis in vivo. Here, we investigate the role of ISCA2 in ccRCC and evaluate its value as a therapeutic target, using a second generation ISCA2 inhibitor - KD061. Methods: To investigate the role of ISCA2 in ccRCC we constitutively overexpressed ISCA2 in ccRCC cells and measured impact on in vitro and in vivo tumor growth, and stained normal human kidney and ccRCC tissue for ISCA2 via IHC. To characterize the mechanism of KD061-induced death, we performed resazurin viability assays and 581/591 BODIPY C11 (lipid peroxidation assays) of KD061 alone or in combination with NAC (anti-oxidant) or liproxstatin (lipid peroxidation inhibitor). We also assessed impact of KD061 on metals uptake using ICP-MS. Impact of KD061 on iron- and stress-activated signaling pathways were evaluated via Western blotting. Anti-tumor efficacy of KD061 was evaluated in mouse models of kidney cancer and tumor ferroptosis was assessed using the MDA assay. Results: ISCA2 was significantly decreased in kidney cancer compared to uninvolved normal kidney. ISCA2 overexpression did not affect in vitro proliferation but significantly decreased tumor growth in vivo. ISCA2 overexpression also promoted resistance to KD061. KD061 induced significant lipid peroxidation that was reversed by co-treatment with NAC or liproxstatin. KD061 treatment resulted in significant increases in metals content and with activation of stress signaling and suppression of HIF-1/2α and a subset of Fe-S client proteins. Daily oral dosing of mice with KD061 was well-tolerated resulting in significant >65% tumor growth inhibition in the RENCA and LVRCC67 syngeneic kidney cancer models, accompanied by increased tumor lipid peroxidation. Conclusions: ISCA2 is decreased in kidney cancer yet continues to play an important role in iron homeostasis. ISCA2 inhibition by KD061 promotes iron dysregulation resulting in iron/metals overload, increased oxidative stress and death via ferroptosis and in decreased HIF-1/2α. KD061 is well tolerated and results in significant tumor growth inhibition in vivo. Thus, KD061 is a promising treatment strategy for kidney cancer. Citation Format: Marin Jones, Yangsook Green, Ethan Reichert, Maria Carolina Ferreira dos Santos, Mei Yee Koh. Targeting ISCA2 and Fe-S assembly to induce ferroptosis in kidney cancer [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Innovations in Kidney Cancer Research: From Molecular Insights to Therapeutic Breakthroughs; 2026 Mar 13-16; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2026;86(5_Suppl_2):Abstract nr A035.