A two-hit model of alcoholic liver disease that exhibits rapid, severe fibrosis.

Alcoholic liver disease (ALD) is responsible for an average of 50.4% and 44.2%of liver disease deaths among males and females respectively. Driven by alcohol misuse, ALD is often reversible by cessation of consumption. However, abstinence programs can have limited success at curtailing abuse, and the loss of life. ALD, therefore, remains a significant clinical challenge. There is a need for effective treatments that prevent or reverse alcohol-induced liver damage to complement or supplant behavioral interventions. Metabolic syndrome, which is disproportionally prevalent in ALD patients, accelerates the progression of ALD and increases liver disease mortality. Current rodent models of ALD unfortunately do not account for the contribution of the western diet to ALD pathology. To address this, we have developed a rodent model of ALD that integrates the impact of the western diet and alcohol; the WASH-diet model. We show here that the WASH diet, either chronically or in small time-restricted bouts, accelerated ALD pathology with severe steatohepatitis, elevated inflammation and increased fibrosis compared to mice receiving chronic alcohol alone. We also validated our WASH-diet model as an in vivo system for testing the efficacy of experimental ALD treatments. The efficacy of the inverse-agonist SR9238, previously shown to inhibit both non-alcohol and alcohol-induced steatohepatitis progression, was conserved in our WASH-diet model. These findings suggested that the WASH-diet may be useful for in vivo pre-clinical assessment of novel therapies.

CRTC1/MAML2 directs a PGC-1α-IGF-1 circuit that confers vulnerability to PPARγ inhibition.

Mucoepidermoid carcinoma (MEC) is a life-threatening salivary gland cancer that is driven primarily by a transcriptional coactivator fusion composed of cyclic AMP-regulated transcriptional coactivator 1 (CRTC1) and mastermind-like 2 (MAML2). The mechanisms by which the chimeric CRTC1/MAML2 (C1/M2) oncoprotein rewires gene expression programs that promote tumorigenesis remain poorly understood. Here, we show that C1/M2 induces transcriptional activation of the non-canonical peroxisome proliferator-activated receptor gamma coactivator-1 alpha (PGC-1α) splice variant PGC-1α4, which regulates peroxisome proliferator-activated receptor gamma (PPARγ)-mediated insulin-like growth factor 1 (IGF-1) expression. This mitogenic transcriptional circuitry is consistent across cell lines and primary tumors. C1/M2-positive tumors exhibit IGF-1 pathway activation, and small-molecule drug screens reveal that tumor cells harboring the fusion gene are selectively sensitive to IGF-1 receptor (IGF-1R) inhibition. Furthermore, this dependence on autocrine regulation of IGF-1 transcription renders MEC cells susceptible to PPARγ inhibition with inverse agonists. These results yield insights into the aberrant coregulatory functions of C1/M2 and identify a specific vulnerability that can be exploited for precision therapy.

LXR-inverse agonism stimulates immune-mediated tumor destruction by enhancing CD8 T-cell activity in triple negative breast cancer.

Triple-negative breast cancer (TNBC) is a highly aggressive subtype that is untreatable with hormonal or HER2-targeted therapies and is also typically unresponsive to checkpoint-blockade immunotherapy. Within the tumor microenvironment dysregulated immune cell metabolism has emerged as a key mechanism of tumor immune-evasion. We have discovered that the Liver-X-Receptors (LXRα and LXRß), nuclear receptors known to regulate lipid metabolism and tumor-immune interaction, are highly activated in TNBC tumor associated myeloid cells. We therefore theorized that inhibiting LXR would induce immune-mediated TNBC-tumor clearance. Here we show that pharmacological inhibition of LXR activity induces tumor destruction primarily through stimulation of CD8+ T-cell cytotoxic activity and mitochondrial metabolism. Our results imply that LXR inverse agonists may be a promising new class of TNBC immunotherapies.

Inhibition of Hepatotoxicity by a LXR Inverse Agonist in a Model of Alcoholic Liver Disease.

Alcohol abuse is a major cause of liver disease and mortality worldwide and is a significant public health issue. Patients with alcoholic liver disease (ALD) have severe hepatic lipid accumulation, inflammation, and fibrosis. Therapies for ALD are very limited and even abstinence from alcohol consumption does not necessarily protect patients from progression of the disease. We sought to evaluate the efficacy of a liver X receptor (LXR) inverse agonist, SR9238, in an animal model of ALD. SR9238 suppresses hepatic lipogenesis, a pathological hallmark of ALD, and we hypothesized that targeting suppression of hepatic metabolic pathways that are activated in ALD may be an effective treatment for the disease. A chronic ethanol diet with or without a final ethanol binge treatment was used to induce ALD in mice. Mice were administered the liver specific LXR inverse agonist SR9238 for 4 weeks after the mice had been maintained on the ethanol diet for 14 days. Mice developed all the hallmarks of advanced ALD demonstrating significant pathophysiology and hepatotoxicity. SR9238 significantly attenuated liver injury and hepatic steatosis and fibrosis was nearly eliminated in SR9238 treated mice. SR9238 treatment reversed the damage associated with chronic ethanol use returning the liver to near normal morphology. These results indicate that inhibiting LXR activity using the inverse agonist has a hepatoprotective effect in rodent models of ALD; thus, this pharmacological approach may be efficacious for treatment of ALD in humans.

Rev-Erb co-regulates muscle regeneration via tethered interaction with the NF-Y cistrome.

OBJECTIVE: The loss of skeletal muscle mass and strength are a central feature of traumatic injury and degenerative myopathies. Unfortunately, pharmacological interventions typically fail to stem the long-term decline in quality of life. Reduced Rev-Erb-mediated gene suppression in cultured C2C12 myoblasts has been shown to stimulate myoblast differentiation. Yet the mechanisms that allow Rev-Erb to pleiotropically inhibit muscle differentiation are not well understood. In this study, we sought to elucidate the role of Rev-Erb in the regulation of muscle differentiation and regeneration in vivo. METHODS: Using Rev-Erbα/ß shRNAs, pharmacological ligands, and Rev-Erbα null and heterozygous mice, we probed the mechanism of Rev-Erbα/ß regulation of muscle differentiation and muscle regeneration. RESULTS: ChIP seq analysis of Rev-Erb in differentiating myoblasts showed that Rev-Erbα did not transcriptionally regulate muscle differentiation through cognate Rev-Erb/ROR-response elements but through possible interaction with the cell fate regulator NF-Y at CCAAT-motifs. Muscle differentiation is stimulated by Rev-Erb release from CCAAT-motifs at promoter and enhancer elements of a number of myogenesis proteins. Partial loss of Rev-Erb expression in mice heterozygous for Rev-Erbα accelerated muscle repair in vivo whereas Rev-Erb knockout mice showed deficiencies in regenerative repair compared to wild type mice. These phenotypic differences between heterozygous and knockout mice were not apparently dependent on MRF induction in response to injury. Similarly, pharmacological disruption of Rev-Erb suppressive activity in injured muscle accelerated regenerative repair in response to acute injury. CONCLUSIONS: Disrupting Rev-Erb activity in injured muscle accelerates regenerative muscle repair/differentiation through transcriptional de-repression of myogenic programs. Rev-Erb, therefore, may be a potent therapeutic target for a myriad of muscular disorders.

Broad Anti-tumor Activity of a Small Molecule that Selectively Targets the Warburg Effect and Lipogenesis.

Malignant cells exhibit aerobic glycolysis (the Warburg effect) and become dependent on de novo lipogenesis, which sustains rapid proliferation and resistance to cellular stress. The nuclear receptor liver-X-receptor (LXR) directly regulates expression of key glycolytic and lipogenic genes. To disrupt these oncogenic metabolism pathways, we designed an LXR inverse agonist SR9243 that induces LXR-corepressor interaction. In cancer cells, SR9243 significantly inhibited the Warburg effect and lipogenesis by reducing glycolytic and lipogenic gene expression. SR9243 induced apoptosis in tumors without inducing weight loss, hepatotoxicity, or inflammation. Our results suggest that LXR inverse agonists may be an effective cancer treatment approach.