Indisulam is an aryl sulfonamide drug with selective anticancer activity. Its mechanism of action and the basis for its selectivity have so far been unknown. Here we show that indisulam promotes the recruitment of RBM39 (RNA binding motif protein 39) to the CUL4-DCAF15 E3 ubiquitin ligase, leading to RBM39 polyubiquitination and proteasomal degradation. Mutations in RBM39 that prevent its recruitment to CUL4-DCAF15 increase RBM39 stability and confer resistance to indisulam's cytotoxicity. RBM39 associates with precursor messenger RNA (pre-mRNA) splicing factors, and inactivation of RBM39 by indisulam causes aberrant pre-mRNA splicing. Many cancer cell lines derived from hematopoietic and lymphoid lineages are sensitive to indisulam, and their sensitivity correlates with DCAF15 expression levels. Two other clinically tested sulfonamides, tasisulam and chloroquinoxaline sulfonamide, share the same mechanism of action as indisulam. We propose that DCAF15 expression may be a useful biomarker to guide clinical trials of this class of drugs, which we refer to as SPLAMs (splicing inhibitor sulfonamides).

Autophagy increases lifespan of model organisms; however, its role in promoting mammalian longevity is less well-established1,2. Here, we report lifespan and healthspan extension in a mouse model with increased basal autophagy. To determine the effects of constitutively increased autophagy on mammalian health, we generated targeted mutant mice with a F121A (Becn1F121A/F121A) mutation in beclin 1 that decreases its interaction with the negative regulator, Bcl-2. We demonstrate that beclin 1/Bcl-2 interaction is disrupted in multiple tissues in Becn1F121A/F121A knock-in (KI) mice in association with higher levels of basal autophagic flux. Compared to wild-type (WT) littermates, the lifespan of both male and female KI mice is significantly increased. The healthspan of the KI mice also improves as aging-related phenotypes are diminished, including age-related renal and cardiac pathological changes and spontaneous tumorigenesis. Moreover, mice deficient in the anti-aging protein, Klotho3, have increased beclin 1/Bcl-2 interaction, decreased autophagy, premature lethality and infertility which are rescued by the beclin 1 F121A mutation. Taken together, our data demonstrate that disruption of the beclin 1/Bcl-2 complex is an effective mechanism to increase autophagy, prevent premature aging, improve healthspan and promote longevity in mammals.

BackgroundVascular calcification is a common feature in patients with chronic kidney disease (CKD). CKD increases serum levels of tumor necrosis factor‐α (TNFα), a critical mediator of vascular calcification. However, the molecular mechanism by which TNFα promotes CKD‐dependent vascular calcification remains obscure. The purpose of the present study was to investigate whether TNFα‐induced vascular calcification in CKD is caused by the endoplasmic reticulum response involving protein kinase RNA‐like endoplasmic reticulum kinase (PERK), eukaryotic initiation factor 2α (eIF2α), activating transcription factor 4 (ATF4), and C/EBP homologous protein (CHOP).Methods and ResultsWe examined the effects of TNFα on the endoplasmic reticulum (ER) stress response of vascular smooth muscle cells (VSMCs). TNFα treatment drastically induced the PERK‐eIF2α‐ATF4‐CHOP axis of the ER stress response in VSMCs. PERK, ATF4, and CHOP shRNA‐mediated knockdowns drastically inhibited mineralization and osteogenesis of VSMCs induced by TNFα. CKD induced by 5/6 nephrectomies activated the PERK‐eIF2α‐ATF4‐CHOP axis of the ER stress response in the aortas of ApoE−/− mice with increased aortic TNFα expression and vascular calcification. Treatment of 5/6 nephrectomized ApoE−/− mice with the TNFα neutralizing antibody or chemical Chaperones reduced aortic PERK‐eIF2α‐ATF4‐CHOP signaling of the ER stress increased by CKD. This resulted in the inhibition of CKD‐dependent vascular calcification.ConclusionsThese results suggest that TNFα induces the PERK‐eIF2α‐ATF4‐CHOP axis of the ER stress response, leading to CKD‐dependent vascular calcification.

Impairment of the autophagy pathway has been observed during the pathogenesis of Alzheimer’s disease (AD), a neurodegenerative disorder characterized by abnormal deposition of extracellular and intracellular amyloid β (Aβ) peptides. Yet the role of autophagy in Aβ production and AD progression is complex. To study whether increased basal autophagy plays a beneficial role in Aβ clearance and cognitive improvement, we developed a novel genetic model to hyperactivate autophagy in vivo. We found that knock-in of a point mutation F121A in the essential autophagy gene Beclin 1/Becn1 in mice significantly reduces the interaction of BECN1 with its inhibitor BCL2, and thus leads to constitutively active autophagy even under non-autophagy-inducing conditions in multiple tissues, including brain. Becn1F121A-mediated autophagy hyperactivation significantly decreases amyloid accumulation, prevents cognitive decline, and restores survival in AD mouse models. Using an immunoisolation method, we found biochemically that Aβ oligomers are autophagic substrates and sequestered inside autophagosomes in the brain of autophagy-hyperactive AD mice. In addition to genetic activation of autophagy by Becn1 gain-of-function, we also found that ML246, a small-molecule autophagy inducer, as well as voluntary exercise, a physiological autophagy inducer, exert similar Becn1-dependent protective effects on Aβ removal and memory in AD mice. Taken together, these results demonstrate that genetically disrupting BECN1-BCL2 binding hyperactivates autophagy in vivo, which sequestrates amyloid oligomers and prevents AD progression. The study establishes new approaches to activate autophagy in the brain, and reveals the important function of Becn1-mediated autophagy hyperactivation in the prevention of AD.