Unraveling the regulatory role of endoplasmic-reticulum-associated degradation in tumor immunity.

During malignant transformation and cancer progression, tumor cells face both intrinsic and extrinsic stress, endoplasmic reticulum (ER) stress in particular. To survive and proliferate, tumor cells use multiple stress response pathways to mitigate ER stress, promoting disease aggression and treatment resistance. Among the stress response pathways is ER-associated degradation (ERAD), which consists of multiple components and steps working together to ensure protein quality and quantity. In addition to its established role in stress responses and tumor cell survival, ERAD has recently been shown to regulate tumor immunity. Here we summarize current knowledge on how ERAD promotes protein degradation, regulates immune cell development and function, participates in antigen presentation, exerts paradoxical roles on tumorigenesis and immunity, and thus impacts current cancer therapy. Collectively, ERAD is a critical protein homeostasis pathway intertwined with cancer development and tumor immunity. Of particular importance is the need to further unveil ERAD's enigmatic roles in tumor immunity to develop effective targeted and combination therapy for successful treatment of cancer.

Base pair opening in a deoxynucleotide duplex containing a cis-syn thymine cyclobutane dimer lesion.

The cis-syn thymine cyclobutane dimer is a DNA photoproduct implicated in skin cancer. We compared the stability of individual base pairs in thymine dimer-containing duplexes to undamaged parent 10-mer duplexes. UV melting thermodynamic measurements, CD spectroscopy, and 2D NOESY NMR spectroscopy confirm that the thymine dimer lesion is locally and moderately destabilizing within an overall B-form duplex conformation. We measured the rates of exchange of individual imino protons by NMR using magnetization transfer from water and determined the equilibrium constant for the opening of each base pair K(op). In the normal duplex K(op) decreases from the frayed ends of the duplex toward the center, such that the central TA pair is the most stable with a K(op) of 8 × 10⁻7. In contrast, base pair opening at the 5'T of the thymine dimer is facile. The 5'T of the dimer has the largest equilibrium constant (K(op) = 3 × 10⁻4) in its duplex, considerably larger than even the frayed penultimate base pairs. Notably, base pairing by the 3'T of the dimer is much more stable than by the 5'T, indicating that the predominant opening mechanism for the thymine dimer lesion is not likely to be flipping out into solution as a single unit. The dimer asymmetrically affects the stability of the duplex in its vicinity, destabilizing base pairing on its 5' side more than on the 3' side. The striking differences in base pair opening between parent and dimer duplexes occur independently of the duplex-single strand melting transitions.

α-Ketoglutarate-Mediated DNA Demethylation Sustains T-Acute Lymphoblastic Leukemia upon TCA Cycle Targeting.

Despite the development of metabolism-based therapy for a variety of malignancies, resistance to single-agent treatment is common due to the metabolic plasticity of cancer cells. Improved understanding of how malignant cells rewire metabolic pathways can guide the rational selection of combination therapy to circumvent drug resistance. Here, we show that human T-ALL cells shift their metabolism from oxidative decarboxylation to reductive carboxylation when the TCA cycle is disrupted. The α-ketoglutarate dehydrogenase complex (KGDHC) in the TCA cycle regulates oxidative decarboxylation by converting α-ketoglutarate (α-KG) to succinyl-CoA, while isocitrate dehydrogenase (IDH) 1 and 2 govern reductive carboxylation. Metabolomics flux analysis of T-ALL reveals enhanced reductive carboxylation upon genetic depletion of the E2 subunit of KGDHC, dihydrolipoamide-succinyl transferase (DLST), mimicking pharmacological inhibition of the complex. Mechanistically, KGDHC dysfunction causes increased demethylation of nuclear DNA by α-KG-dependent dioxygenases (e.g., TET demethylases), leading to increased production of both IDH1 and 2. Consequently, dual pharmacologic inhibition of the TCA cycle and TET demethylases demonstrates additive efficacy in reducing the tumor burden in zebrafish xenografts. These findings provide mechanistic insights into how T-ALL develops resistance to drugs targeting the TCA cycle and therapeutic strategies to overcome this resistance.

Cross organelle stress response disruption promotes gentamicin-induced proteotoxicity.

Gentamicin is a nephrotoxic antibiotic that causes acute kidney injury (AKI) primarily by targeting the proximal tubule epithelial cell. The development of an effective therapy for gentamicin-induced renal cell injury is limited by incomplete mechanistic insight. To address this challenge, we propose that RNAi signal pathway screening could identify a unifying mechanism of gentamicin-induced cell injury and suggest a therapeutic strategy to ameliorate it. Computational analysis of RNAi signal screens in gentamicin-exposed human proximal tubule cells suggested the cross-organelle stress response (CORE), the unfolded protein response (UPR), and cell chaperones as key targets of gentamicin-induced injury. To test this hypothesis, we assessed the effect of gentamicin on the CORE, UPR, and cell chaperone function, and tested the therapeutic efficacy of enhancing cell chaperone content. Early gentamicin exposure disrupted the CORE, evidenced by a rise in the ATP:ADP ratio, mitochondrial-specific H2O2 accumulation, Drp-1-mediated mitochondrial fragmentation, and endoplasmic reticulum-mitochondrial dissociation. CORE disruption preceded measurable increases in whole-cell oxidative stress, misfolded protein content, transcriptional UPR activation, and its untoward downstream effects: CHOP expression, PARP cleavage, and cell death. Geranylgeranylacetone, a therapeutic that increases cell chaperone content, prevented mitochondrial H2O2 accumulation, preserved the CORE, reduced the burden of misfolded proteins and CHOP expression, and significantly improved survival in gentamicin-exposed cells. We identify CORE disruption as an early and remediable cause of gentamicin proteotoxicity that precedes downstream UPR activation and cell death. Preserving the CORE significantly improves renal cell survival likely by reducing organelle-specific proteotoxicity during gentamicin exposure.