First-in-class TMBIM6 therapeutics: Overcoming limitations in current oncology therapies through ERAD-II induced paraptosis.

e15120 Background: Despite advances in cancer treatment, significant challenges remain, such as immune evasion, chemoresistance, and reliance on apoptotic pathways. TMBIM6, a stress-responsive intracellular protein, has been well researched for its role in many diseases/injuries. Within cancer, TMBIM6 plays a key role in tumorigenesis, chemoresistance, metastasis and cancer survival. TMBIM6 is only ‘activated’ by IntraCellular Environmental (ICE) changes e.g., high cytosolic; calcium; ROS; acidification; etc. In cancer, ICE changes are a byproduct of the disease and not essential for pathogenesis. We developed first-in-class small molecules (SMQs) that modulate TMBIM6 inducing rapid cancer cell death with no measured/observed toxicity. Methods: SMQ effect was measured on commercial and PDX cell lines that represent all sub-types of breast, ovarian, lung, colorectal, and uterine cancers. We assessed SMQ impact on cell viability, morphological characteristics, and intracellular changes. Mitochondrial and cytosolic ROS, calcium levels, and lysosomal activity were quantified using live-cell imaging and biochemical assays. Genetic knockdown and TMBIM6 mutational analysis were conducted to evaluate their impact in SMQ effects. In vivo we analyzed tumor volume, weight, and toxicity. Results: In vitro : SMQ induced > 95% cancer cell death within 72-96 hours of treatment (p < 0.0001). This effect was independent of cancer genotype, phenotype, or TMBIM6 expression levels but required functional TMBIM6 and TMBIM6 environmental detection site (D213). In vivo : Breast (Triple Negative and Luminal A sub-type) and Ovarian (HSOC) mouse models showed > 95% reduction in tumor size and volume within 25 days with no impact on mouse body weight, size, or behavior. Toxicity assessment, from blood samples and IHC organ staining, showed no measured or observable toxicity Mechanism: SMQs induced paraptotic cell death, characterized by extensive ER vacuolation, mitochondrial membrane permeabilization, and elevated cytosolic ROS/calcium. ROS and calcium scavengers inhibited paraptosis, confirming their critical roles in the mechanism. Lysosomal activation and non-canonical ER-associated degradation pathways were identified as central to this mechanism. Importantly, SMQ had no impact on non-cancerous cells or healthy tissue, even at high doses. Conclusions: Our studies show activating TMBIM6 induces paraptosis — a non-apoptotic, immune-silent form of programmed cell death. This effect was universal across all cancers tested, irrespective of TMBIM6 expression level, genotype and phenotype. TMBIM6 is potentially a groundbreaking therapeutic target, and SMQs could be a safe therapeutic option for solid tumors and resistant malignancies. Our findings strongly advocate for advancing SMQs into clinical development as an adjunct or alternative to current oncology treatment strategies.