Darville, L. | bioRxiv Science

bioRxiv2026

TAp63-regulated oncogenic long non-coding RNA-8 ( TROLL-8 ) regulates human breast cancer progression through CPT1A-mediated fatty acid oxidation

Metabolic reprogramming is a crucial hallmark of cancer, supporting tumor growth and adaptation to cellular stress. Although fatty acid oxidation (FAO) has emerged as an important regulator in cancer, the mechanisms that control the FAO machinery remain poorly understood. Here, we demonstrate that the TAp63-regulated long non-coding RNA TROLL-8 is a key regulator of FAO in breast cancer. Using metabolomics and protein microarray assays followed by immunoprecipitation-mass spectrometry, we mechanistically demonstrate that TROLL-8 binds the FAO enzyme CPT1A and promotes the formation of a complex with ACSL1 and VDAC1, thereby enabling efficient fatty acid processing. Loss of TROLL-8 destabilizes this complex, leading to impaired FAO, decreased metabolic fitness, and suppression of tumorigenic phenotypes, such as anchorage-independent growth. Notably, higher levels of CPT1A and VDAC1 are associated with worse survival in breast cancer patients. Given the emerging role of CPT1A in therapy resistance, these findings suggest that targeting TROLL-8 could be a promising approach to selectively disrupt the hyperactive FAO machinery in breast cancers and other tumor types that rely on FAO for their progression.

Cancer Biology↗

bioRxiv2026

C-terminus CD28 phosphorylation (Y218) modulates IL-2 secretion and antitumor effect of CAR-T cells

CD28 is a co-stimulatory component of several second-generation chimeric antigen receptor (CAR)-T cells, providing signals essential for T cell proliferation, survival, and cytokine secretion. However, the specific contribution of individual CD28 intracellular motifs to CAR-T cell function remains incompletely understood. Here, we identify tyrosine 218 (Y218) in the CD28 cytoplasmic domain as a critical regulatory site, and demonstrate that its phosphorylation is essential for optimal CAR-T cell activity. Using a 218F mutant, we show that loss of Y218 phosphorylation leads to impaired IL-2 production and abrogates antitumor efficacy. Transcriptomic profiling of 218F CAR-T cells revealed increased expression of IL-17A, IL-17F, and related cytokines, suggesting a shift toward a pro-inflammatory Th17-like phenotype that may contribute to dysfunction. Mechanistically, we demonstrate that the interleukin-2-inducible T-cell kinase, ITK, mediates Y218 phosphorylation. To further understand the role of this kinase, we engineered a novel CAR incorporating an ITK-binding motif (PYRP), which enhances ITK recruitment, increases Y218 phosphorylation, and boosts IL-2 secretion, and improves anti-tumor efficacy in vivo. Our findings underscore the functional relevance of Y218 phosphorylation in modulating CAR-T cell fate and reveal a strategy to fine-tune CAR signaling through targeted kinase recruitment to enhance therapeutic efficacy.

immunology↗