Suppression of Ah Receptor (AhR) increases the aggressiveness of TNBC cells and 11-Cl-BBQ-activated AhR inhibits their growth.

Triple-negative breast cancer (TNBC) represents around 15% of the 2.26 million breast cancers diagnosed worldwide annually and has the worst outcome. Despite recent therapeutic advances, there remains a lack of targeted therapies for this breast cancer subtype. The aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor with biological roles in regulating development, xenobiotic metabolism, cell cycle progression and cell death. AhR activation by select ligands can promote tumor suppression in multiple cancer types. AhR can negatively regulate the activity of different oncogenic signaling pathways and can directly upregulate tumor suppressor genes such as p27Kip1. To determine the role of AhR in TNBC, we generated AhR-deficient cancer cells and investigated the impact of AhR loss on TNBC cell growth phenotypes. We found that AhR-deficient MDA-MB-468 TNBC cells have increased proliferation and formed significantly more colonies compared to AhR expressing cells. These cells without AhR expression grew aggressively in vivo. To determine the molecular targets driving this phenotype, we performed transcriptomic profiling in AhR expressing and AhR knockout MDA-MB-468 cells and identified tyrosine receptor kinases, as well as other genes involved in proliferation, survival and clonogenicity that are repressed by AhR. In order to determine therapeutic targeting of AhR in TNBC, we investigated the anti-cancer effects of the novel AhR ligand 11-chloro-7H-benzimidazo[2,1-a]benzo[de]iso-quinolin-7-one (11-Cl-BBQ), which belongs to a class of high affinity, rapidly metabolized AhR ligands called benzimidazoisoquinolines (BBQs). 11-Cl-BBQ induced AhR-dependent cancer cell-selective growth inhibition and strongly inhibited colony formation in TNBC cells.

Necroptosis inhibition counteracts neurodegeneration, memory decline, and key hallmarks of aging, promoting brain rejuvenation.

Age is the main risk factor for the development of neurodegenerative diseases. In the aged brain, axonal degeneration is an early pathological event, preceding neuronal dysfunction, and cognitive disabilities in humans, primates, rodents, and invertebrates. Necroptosis mediates degeneration of injured axons, but whether necroptosis triggers neurodegeneration and cognitive impairment along aging is unknown. Here, we show that the loss of the necroptotic effector Mlkl was sufficient to delay age-associated axonal degeneration and neuroinflammation, protecting against decreased synaptic transmission and memory decline in aged mice. Moreover, short-term pharmacologic inhibition of necroptosis targeting RIPK3 in aged mice, reverted structural and functional hippocampal impairment, both at the electrophysiological and behavioral level. Finally, a quantitative proteomic analysis revealed that necroptosis inhibition leads to an overall improvement of the aged hippocampal proteome, including a subclass of molecular biofunctions associated with brain rejuvenation, such as long-term potentiation and synaptic plasticity. Our results demonstrate that necroptosis contributes to age-dependent brain degeneration, disturbing hippocampal neuronal connectivity, and cognitive function. Therefore, necroptosis inhibition constitutes a potential geroprotective strategy to treat age-related disabilities associated with memory impairment and cognitive decline.

Aberrant cortical spine dynamics after concussive injury are reversed by integrated stress response inhibition.

Traumatic brain injury (TBI) is a leading cause of long-term neurological disability in the world and the strongest environmental risk factor for the development of dementia. Even mild TBI (resulting from concussive injuries) is associated with a greater than twofold increase in the risk of dementia onset. Little is known about the cellular mechanisms responsible for the progression of long-lasting cognitive deficits. The integrated stress response (ISR), a phylogenetically conserved pathway involved in the cellular response to stress, is activated after TBI, and inhibition of the ISR-even weeks after injury-can reverse behavioral and cognitive deficits. However, the cellular mechanisms by which ISR inhibition restores cognition are unknown. Here, we used longitudinal two-photon imaging in vivo after concussive injury in mice to study dendritic spine dynamics in the parietal cortex, a brain region involved in working memory. Concussive injury profoundly altered spine dynamics measured up to a month after injury. Strikingly, brief pharmacological treatment with the drug-like small-molecule ISR inhibitor ISRIB entirely reversed structural changes measured in the parietal cortex and the associated working memory deficits. Thus, both neural and cognitive consequences of concussive injury are mediated in part by activation of the ISR and can be corrected by its inhibition. These findings suggest that targeting ISR activation could serve as a promising approach to the clinical treatment of chronic cognitive deficits after TBI.

Correction of eIF2-dependent defects in brain protein synthesis, synaptic plasticity, and memory in mouse models of Alzheimer's disease.

Neuronal protein synthesis is essential for long-term memory consolidation, and its dysregulation is implicated in various neurodegenerative disorders, including Alzheimer's disease (AD). Cellular stress triggers the activation of protein kinases that converge on the phosphorylation of eukaryotic translation initiation factor 2α (eIF2α), which attenuates mRNA translation. This translational inhibition is one aspect of the integrated stress response (ISR). We found that postmortem brain tissue from AD patients showed increased phosphorylation of eIF2α and reduced abundance of eIF2B, another key component of the translation initiation complex. Systemic administration of the small-molecule compound ISRIB (which blocks the ISR downstream of phosphorylated eIF2α) rescued protein synthesis in the hippocampus, measures of synaptic plasticity, and performance on memory-associated behavior tests in wild-type mice cotreated with salubrinal (which inhibits translation by inducing eIF2α phosphorylation) and in both ß-amyloid-treated and transgenic AD model mice. Thus, attenuating the ISR downstream of phosphorylated eIF2α may restore hippocampal protein synthesis and delay cognitive decline in AD patients.

Small molecule cognitive enhancer reverses age-related memory decline in mice.

With increased life expectancy, age-associated cognitive decline becomes a growing concern, even in the absence of recognizable neurodegenerative disease. The integrated stress response (ISR) is activated during aging and contributes to age-related brain phenotypes. We demonstrate that treatment with the drug-like small-molecule ISR inhibitor ISRIB reverses ISR activation in the brain, as indicated by decreased levels of activating transcription factor 4 (ATF4) and phosphorylated eukaryotic translation initiation factor eIF2. Furthermore, ISRIB treatment reverses spatial memory deficits and ameliorates working memory in old mice. At the cellular level in the hippocampus, ISR inhibition (i) rescues intrinsic neuronal electrophysiological properties, (ii) restores spine density and (iii) reduces immune profiles, specifically interferon and T cell-mediated responses. Thus, pharmacological interference with the ISR emerges as a promising intervention strategy for combating age-related cognitive decline in otherwise healthy individuals.

Integrated Stress Response Inhibitor Reverses Sex-Dependent Behavioral and Cell-Specific Deficits after Mild Repetitive Head Trauma.

Mild repetitive traumatic brain injury (rTBI) induces chronic behavioral and cognitive alterations and increases the risk for dementia. Currently, there are no therapeutic strategies to prevent or mitigate chronic deficits associated with rTBI. Previously we developed an animal model of rTBI that recapitulates the cognitive and behavioral deficits observed in humans. We now report that rTBI results in an increase in risk-taking behavior in male but not female mice. This behavioral phenotype is associated with chronic activation of the integrated stress response and cell-specific synaptic alterations in the type A subtype of layer V pyramidal neurons in the medial prefrontal cortex. Strikingly, by briefly treating animals weeks after injury with ISRIB, a selective inhibitor of the integrated stress response (ISR), we (1) relieve ISR activation, (2) reverse the increased risk-taking behavioral phenotype and maintain this reversal, and (3) restore cell-specific synaptic function in the affected mice. Our results indicate that targeting the ISR even at late time points after injury can permanently reverse behavioral changes. As such, pharmacological inhibition of the ISR emerges as a promising avenue to combat rTBI-induced behavioral dysfunction.

Dual Inhibition of Bruton's Tyrosine Kinase and Phosphoinositide-3-Kinase p110δ as a Therapeutic Approach to Treat Non-Hodgkin's B Cell Malignancies.

Although new targeted therapies, such as ibrutinib and idelalisib, have made a large impact on non-Hodgkin's lymphoma (NHL) patients, the disease is often fatal because patients are initially resistant to these targeted therapies, or because they eventually develop resistance. New drugs and treatments are necessary for these patients. One attractive approach is to inhibit multiple parallel pathways that drive the growth of these hematologic tumors, possibly prolonging the duration of the response and reducing resistance. Early clinical trials have tested this approach by dosing two drugs in combination in NHL patients. We discovered a single molecule, MDVN1003 (1-(5-amino-2,3-dihydro-1H-inden-2-yl)-3-(8-fluoro-3,4-dihydro-2H-benzo[b][1,4]oxazin-6-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine), that inhibits Bruton's tyrosine kinase and phosphatidylinositol-3-kinase δ, two proteins regulated by the B cell receptor that drive the growth of many NHLs. In this report, we show that this dual inhibitor prevents the activation of B cells and inhibits the phosphorylation of protein kinase B and extracellular signal-regulated kinase 1/2, two downstream mediators that are important for this process. Additionally, MDVN1003 induces cell death in a B cell lymphoma cell line but not in an irrelevant erythroblast cell line. Importantly, we found that this orally bioavailable dual inhibitor reduced tumor growth in a B cell lymphoma xenograft model more effectively than either ibrutinib or idelalisib. Taken together, these results suggest that dual inhibition of these two key pathways by a single molecule could be a viable approach for treatment of NHL patients.

Purinergic Signaling as a Regulator of Th17 Cell Plasticity.

T helper type 17 (Th17) lymphocytes, characterized by the production of interleukin-17 and other pro-inflammatory cytokines, are present in intestinal lamina propria and have been described as important players driving intestinal inflammation. Recent evidence, supporting the notion of a functional and phenotypic instability of Th17 cells, has shown that Th17 differentiate into type 1 regulatory (Tr1) T cells during the resolution of intestinal inflammation. Moreover, it has been suggested that the expression of CD39 ectonucleotidase endows Th17 cells with immunosuppressive properties. However, the exact role of CD39 ectonucleotidase in Th17 cells has not been studied in the context of intestinal inflammation. Here we show that Th17 cells expressing CD39 ectonucleotidase can hydrolyze ATP and survive to ATP-induced cell death. Moreover, in vitro-generated Th17 cells expressing the CD39 ectonucleotidase produce IL-10 and are less pathogenic than CD39 negative Th17 cells in a model of experimental colitis in Rag-/- mice. Remarkably, we show that CD39 activity regulates the conversion of Th17 cells to IL-10-producing cells in vitro, which is abrogated in the presence of ATP and the CD39-specific inhibitor ARL67156. All these data suggest that CD39 expression by Th17 cells allows the depletion of ATP and is crucial for IL-10 production and survival during the resolution of intestinal inflammation.