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α2δ-1 (encoded by theCacna2d1gene) is a newly discovered NMDA receptor-interacting protein and is the therapeutic target of gabapentinoids (e.g., gabapentin and pregabalin) frequently used for treating patients with neuropathic pain. Nerve injury causes sustained α2δ-1 upregulation in the dorsal root ganglion (DRG), which promotes NMDA receptor synaptic trafficking and activation in the spinal dorsal horn, a hallmark of chronic neuropathic pain. However, little is known about how nerve injury initiates and maintains the high expression level of α2δ-1 to sustain chronic pain. Here, we show that nerve injury caused histone hyperacetylation and diminished enrichment of histone deacetylase-2 (HDAC2), but not HDAC3, at theCacna2d1promoter in the DRG. Strikingly,Hdac2knockdown or conditional knockout in DRG neurons in male and female mice consistently induced long-lasting mechanical pain hypersensitivity, which was readily reversed by blocking NMDA receptors, inhibiting α2δ-1 with gabapentin or disrupting the α2δ-1–NMDA receptor interaction at the spinal cord level.Hdac2deletion in DRG neurons increased histone acetylation levels at theCacna2d1promoter, upregulated α2δ-1 in the DRG, and potentiated α2δ-1–dependent NMDA receptor activity at primary afferent central terminals in the spinal dorsal horn. Correspondingly,Hdac2knockdown-induced pain hypersensitivity was blunted inCacna2d1knockout mice. Thus, our findings reveal that HDAC2 functions as a pivotal transcriptional repressor of neuropathic pain via constitutively suppressing α2δ-1 expression and ensuing presynaptic NMDA receptor activity in the spinal cord. HDAC2 enrichment levels at theCacna2d1promoter in DRG neurons constitute a unique epigenetic mechanism that governs acute-to-chronic pain transition.SIGNIFICANCE STATEMENTExcess α2δ-1 proteins produced after nerve injury directly interact with glutamate NMDA receptors to potentiate synaptic NMDA receptor activity in the spinal cord, a prominent mechanism of nerve pain. Because α2δ-1 upregulation after nerve injury is long lasting, gabapentinoids relieve pain symptoms only temporarily. Our study demonstrates for the first time the unexpected role of intrinsic HDAC2 activity at the α2δ-1 gene promoter in limiting α2δ-1 gene transcription, NMDA receptor-dependent synaptic plasticity, and chronic pain development after nerve injury. These findings challenge the prevailing view about the role of general HDAC activity in promoting chronic pain. Restoring the repressive HDAC2 function and/or reducing histone acetylation at the α2δ-1 gene promoter in primary sensory neurons could lead to long-lasting relief of nerve pain.

Activating mutations in KRAS (KRAS*) are present in nearly all pancreatic ductal adenocarcinoma (PDAC) cases and critical for tumor maintenance. By using an inducible KRAS* PDAC mouse model, we identified a deubiquitinase USP21-driven resistance mechanism to anti-KRAS* therapy. USP21 promotes KRAS*-independent tumor growth via its regulation of MARK3-induced macropinocytosis, which serves to maintain intracellular amino acid levels for anabolic growth. The USP21-mediated KRAS* bypass, coupled with the frequent amplification of USP21 in human PDAC tumors, encourages the assessment of USP21 as a novel drug target as well as a potential parameter that may affect responsiveness to emergent anti-KRAS* therapy.

Radiotherapy (RT) of colorectal cancer (CRC) can prime adaptive immunity against tumor-associated antigen (TAA)–expressing CRC cells systemically. However, abscopal tumor remissions are extremely rare, and the postirradiation immune escape mechanisms in CRC remain elusive. Here, we found that irradiated CRC cells used ATR-mediated DNA repair signaling pathway to up-regulate both CD47 and PD-L1, which through engagement of SIRPα and PD-1, respectively, prevented phagocytosis by antigen-presenting cells and thereby limited TAA cross-presentation and innate immune activation. This postirradiation CD47 and PD-L1 up-regulation was observed across various human solid tumor cells. Concordantly, rectal cancer patients with poor responses to neoadjuvant RT exhibited significantly elevated postirradiation CD47 levels. The combination of RT, anti-SIRPα, and anti–PD-1 reversed adaptive immune resistance and drove efficient TAA cross-presentation, resulting in robust TAA-specific CD8 T cell priming, functional activation of T effectors, and increased T cell clonality and clonal diversity. We observed significantly higher complete response rates to RT/anti-SIRPα/anti–PD-1 in both irradiated and abscopal tumors and prolonged survival in three distinct murine CRC models, including a cecal orthotopic model. The efficacy of triple combination therapy was STING dependent as knockout animals lost most benefit of adding anti-SIRPα and anti–PD-1 to RT. Despite activation across the myeloid stroma, the enhanced dendritic cell function accounts for most improvements in CD8 T cell priming. These data suggest ATR-mediated CD47 and PD-L1 up-regulation as a key mechanism restraining radiation-induced immune priming. RT combined with SIRPα and PD-1 blockade promotes robust antitumor immune priming, leading to systemic tumor regressions.

Inhibiting glycolysis remains an aspirational approach for the treatment of cancer. We have previously identified a subset of cancers harbouring homozygous deletion of the glycolytic enzyme enolase (ENO1) that have exceptional sensitivity to inhibition of its redundant paralogue, ENO2, through a therapeutic strategy known as collateral lethality. Here, we show that a small-molecule enolase inhibitor, POMHEX, can selectively kill ENO1-deleted glioma cells at low-nanomolar concentrations and eradicate intracranial orthotopic ENO1-deleted tumours in mice at doses well-tolerated in non-human primates. Our data provide an in vivo proof of principle of the power of collateral lethality in precision oncology and demonstrate the utility of POMHEX for glycolysis inhibition with potential use across a range of therapeutic settings.

Tumor-associated macrophages (TAMs) are correlated with the progression of prostatic adenocarcinoma (PCa). The mechanistic basis of this correlation and therapeutic strategies to target TAMs in PCa remain poorly defined. Here, single-cell RNA sequencing was used to profile the transcriptional landscape of TAMs in human PCa, leading to identification of a subset of macrophages characterized by dysregulation in transcriptional pathways associated with lipid metabolism. This subset of TAMs correlates positively with PCa progression and shorter disease-free survival and is characterized by an accumulation of lipids that is dependent on Marco. Mechanistically, cancer cell–derived IL-1β enhances Marco expression on macrophages, and reciprocally, cancer cell migration is promoted by CCL6 released by lipid-loaded TAMs. Moreover, administration of a high-fat diet to tumor-bearing mice raises the abundance of lipid-loaded TAMs. Finally, targeting lipid accumulation by Marco blockade hinders tumor growth and invasiveness and improves the efficacy of chemotherapy in models of PCa, pointing to combinatorial strategies that may influence patient outcomes.

Inflammatory bowel disease (IBD) is a chronic inflammatory condition driven by diverse genetic and nongenetic programs that converge to disrupt immune homeostasis in the intestine. We have reported that, in murine intestinal epithelium with telomere dysfunction, DNA damage-induced activation of ataxia-telangiectasia mutated (ATM) results in ATM-mediated phosphorylation and activation of the YAP1 transcriptional coactivator, which in turn up-regulates pro-IL-18, a pivotal immune regulator in IBD pathogenesis. Moreover, individuals with germline defects in telomere maintenance genes experience increased occurrence of intestinal inflammation and show activation of the ATM/YAP1/pro-IL-18 pathway in the intestinal epithelium. Here, we sought to determine the relevance of the ATM/YAP1/pro-IL-18 pathway as a potential driver of IBD, particularly older-onset IBD. Analysis of intestinal biopsy specimens and organoids from older-onset IBD patients documented the presence of telomere dysfunction and activation of the ATM/YAP1/precursor of interleukin 18 (pro-IL-18) pathway in the intestinal epithelium. Employing intestinal organoids from healthy individuals, we demonstrated that experimental induction of telomere dysfunction activates this inflammatory pathway. In organoid models from ulcerative colitis and Crohn's disease patients, pharmacological interventions of telomerase reactivation, suppression of DNA damage signaling, or YAP1 inhibition reduced pro-IL-18 production. Together, these findings support a model wherein telomere dysfunction in the intestinal epithelium can initiate the inflammatory process in IBD, pointing to therapeutic interventions for this disease.

The ubiquitin-specific protease (USP) family is the largest group of cysteine proteases. Cancer genomic analysis identified frequent amplification of USP21 (22%) in human pancreatic ductal adenocarcinoma (PDAC). USP21 overexpression correlates with human PDAC progression, and enforced expression of USP21 accelerates murine PDAC tumor growth and drives PanIN to PDAC progression in immortalized human pancreatic ductal cells. Conversely, depletion of USP21 impairs PDAC tumor growth. Mechanistically, USP21 deubiquitinates and stabilizes the TCF/LEF transcription factor TCF7, which promotes cancer cell stemness. Our work identifies and validates USP21 as a PDAC oncogene, providing a potential druggable target for this intractable disease.

Multiple myeloma is the second most frequent hematological cancer after lymphoma and remains an incurable disease. The pervasive support provided by the bone marrow microenvironment to myeloma cells is crucial for their survival. Here, an unbiased assessment of receptor tyrosine kinases overexpressed in myeloma identified ROR2, a receptor for the WNT noncanonical pathway, as highly expressed in myeloma cells. Its ligand, WNT5A is the most abundant growth factor in the bone marrow of myeloma patients. ROR2 mediates myeloma cells interactions with the surrounding bone marrow and its depletion resulted in detachment of myeloma cells from their niche in an in vivo model, triggering apoptosis and thus markedly delaying disease progression. Using in vitro and ex vivo 3D-culture systems, ROR2 was shown to exert a pivotal role in the adhesion of cancer cells to the microenvironment. Genomic studies revealed that the pathways mostly deregulated by ROR2 overexpression were PI3K/AKT and mTOR. Treatment of cells with specific PI3K inhibitors already used in the clinic reduced myeloma cell adhesion to the bone marrow. Together, our findings support the view that ROR2 and its downstream targets represent a novel therapeutic strategy for the large subgroup of MM patients whose cancer cells show ROR2 overexpression.

Pancreatic ductal adenocarcinoma (PDAC) remains recalcitrant to all forms of cancer treatment and carries a five-year survival rate of only 8%1. Inhibition of oncogenic KRAS (hereafter KRAS*), the earliest lesion in disease development that is present in more than 90% of PDACs, and its signalling surrogates has yielded encouraging preclinical results with experimental agents2–4. However, KRAS*-independent disease recurrence following genetic extinction of Kras* in mouse models anticipates the need for co-extinction strategies5,6. Multiple oncogenic processes are initiated at the cell surface, where KRAS* physically and functionally interacts to direct signalling that is essential for malignant transformation and tumour maintenance. Insights into the complexity of the functional cell-surface-protein repertoire (surfaceome) have been technologically limited until recently and—in the case of PDAC—the genetic control of the function and composition of the PDAC surfaceome in the context of KRAS* signalling remains largely unknown. Here we develop an unbiased, functional target-discovery platform to query KRAS*-dependent changes of the PDAC surfaceome, which reveals syndecan 1 (SDC1, also known as CD138) as a protein that is upregulated at the cell surface by KRAS*. Localization of SDC1 at the cell surface—where it regulates macropinocytosis, an essential metabolic pathway that fuels PDAC cell growth—is essential for disease maintenance and progression. Thus, our study forges a mechanistic link between KRAS* signalling and a targetable molecule driving nutrient salvage pathways in PDAC and validates oncogene-driven surfaceome annotation as a strategy to identify cancer-specific vulnerabilities. In an inducible mouse model of pancreatic ductal adenocarcinoma, the signalling defect that underlies 90% of these tumours causes increased cell-surface expression of syndecan 1, leading to misregulation of macropinocytosis, and linking the defective signalling with nutrient-salvage pathways.

Summary: The plasticity of a preexisting regulatory circuit compromises the effectiveness of targeted therapies, and leveraging genetic vulnerabilities in cancer cells may overcome such adaptations. Hereditary leiomyomatosis renal cell carcinoma (HLRCC) is characterized by oxidative phosphorylation (OXPHOS) deficiency caused by fumarate hydratase (FH) nullizyogosity. To identify metabolic genes that are synthetically lethal with OXPHOS deficiency, we conducted a genetic loss-of-function screen and found that phosphogluconate dehydrogenase (PGD) inhibition robustly blocks the proliferation of FH mutant cancer cells both in vitro and in vivo. Mechanistically, PGD inhibition blocks glycolysis, suppresses reductive carboxylation of glutamine, and increases the NADP+/NADPH ratio to disrupt redox homeostasis. Furthermore, in the OXPHOS-proficient context, blocking OXPHOS using the small-molecule inhibitor IACS-010759 enhances sensitivity to PGD inhibition in vitro and in vivo. Together, our study reveals a dependency on PGD in OXPHOS-deficient tumors that might inform therapeutic intervention in specific patient populations. : Loss-of-function genetics screen reveals a synthetically lethal interaction between OXPHOS inhibition and phosphogluconate dehydrogenase (PGD) inactivation. Sun et al. provide an example of targeting tumor metabolism in a genetically predefined context to maximize therapeutic impact and propose PGD as a therapeutic target for fumarate hydratase-deficient HLRCC. Keywords: synthetic lethality, PGD, OXPHOS, tumor metabolism, metabolic vulnerability, fumarate hydratase, redox homeostasis, functional genomics, hereditary leiomyomatosis renal cell carcinoma, pentose phosphate pathway

The application of single-cell RNA sequencing platforms has generated notable insights into the heterogeneity underlying pancreatic ductal adenocarcinoma (PDAC), encompassing both the neoplastic compartment and the tumour microenvironment. In this Comment, we discuss the most pertinent findings gleaned from both mouse models and human PDAC samples, as well as future opportunities.

The insulin-like growth factor 1 receptor (IGF1R) and phosphoinositide 3-kinase p110α (PI3K) are critical regulators of exercise-induced physiological cardiac hypertrophy and provide protection in experimental models of pathological remodeling and heart failure. Forkhead box class O1 (FoxO1) is a transcription factor that regulates cardiomyocyte hypertrophy downstream of IGF1R/PI3K activation in vitro, but its role in physiological hypertrophy in vivo was unknown. We generated cardiomyocyte-specific FoxO1 knockout (cKO) mice and assessed the phenotype under basal conditions and settings of physiological hypertrophy induced by 1) swim training or 2) cardiac-specific transgenic expression of constitutively active PI3K (caPI3KTg+). Under basal conditions, male and female cKO mice displayed mild interstitial fibrosis compared with control (CON) littermates, but no other signs of cardiac pathology were present. In response to exercise training, female CON mice displayed an increase (∼21%) in heart weight normalized to tibia length vs. untrained mice. Exercise-induced hypertrophy was blunted in cKO mice. Exercise increased cardiac Akt phosphorylation and IGF1R expression but was comparable between genotypes. However, differences in Foxo3a, Hsp70, and autophagy markers were identified in hearts of exercised cKO mice. Deletion of FoxO1 did not reduce cardiac hypertrophy in male or female caPI3KTg+ mice. Cardiac Akt and FoxO1 protein expressions were significantly reduced in hearts of caPI3KTg+ mice, which may represent a negative feedback mechanism from chronic caPI3K, and negate any further effect of reducing FoxO1 in the cKO. In summary, FoxO1 contributes to exercise-induced hypertrophy. This has important implications when one is considering FoxO1 as a target for treating the diseased heart.NEW & NOTEWORTHY Regulators of exercise-induced physiological cardiac hypertrophy and protection are considered promising targets for the treatment of heart failure. Unlike pathological hypertrophy, the transcriptional regulation of physiological hypertrophy has remained largely elusive. To our knowledge, this is the first study to show that the transcription factor FoxO1 is a critical mediator of exercise-induced cardiac hypertrophy. Given that exercise-induced hypertrophy is protective, this finding has important implications when one is considering FoxO1 as a target for treating the diseased heart.

The escalating social and economic burden of an aging world population has placed aging research at center stage. The hallmarks of aging comprise diverse molecular mechanisms and cellular systems that are interrelated and act in concert to drive the aging process. Here, through the lens of telomere biology, we examine how telomere dysfunction may amplify or drive molecular biological processes underlying each hallmark of aging and contribute to the development of age-related diseases such as neurodegeneration and cancer. The intimate link of telomeres to aging hallmarks informs preventive and therapeutic interventions designed to attenuate aging itself and reduce the incidence of age-associated diseases.

Germline telomere maintenance defects are associated with an increased incidence of inflammatory diseases in humans, yet whether and how telomere dysfunction causes inflammation are not known. Here, we show that telomere dysfunction drives pATM/c-ABL-mediated activation of the YAP1 transcription factor, up-regulating the major pro-inflammatory factor, pro-IL-18. The colonic microbiome stimulates cytosolic receptors activating caspase-1 which cleaves pro-IL-18 into mature IL-18, leading to recruitment of interferon (IFN)-γ-secreting T cells and intestinal inflammation. Correspondingly, patients with germline telomere maintenance defects exhibit DNA damage (γH2AX) signaling together with elevated YAP1 and IL-18 expression. In mice with telomere dysfunction, telomerase reactivation in the intestinal epithelium or pharmacological inhibition of ATM, YAP1, or caspase-1 as well as antibiotic treatment, dramatically reduces IL-18 and intestinal inflammation. Thus, telomere dysfunction-induced activation of the ATM-YAP1-pro-IL-18 pathway in epithelium is a key instigator of tissue inflammation., How telomere dysfunction is directly linked to inflammation in humans is currently unclear. Here the authors reveal that telomere dysfunction drives activation of the YAP1 transcription factor, up-regulating the pro inflammatory factor, pro-IL-18 thus revealing a link between telomere dysfunction and initiation of intestinal inflammation.

In this issue of Cancer Cell, Aftab et al. identify a pro-inflammatory cytokine, IL-33, that is released as a chemoattractant for type 2 immune cells in response to the intratumoral mycobiome. Depletion of fungi or deletion of IL-33 in cancer cells significantly decreases pancreatic ductal adenocarcinoma (PDAC) tumor progression and increases survival.

Dysregulated ROR-γt-mediated IL-17 transcription is central to the pathogenesis of several inflammatory disorders, yet the molecular mechanisms that govern the transcription factor activity of ROR-γt in the regulation of IL-17 are not fully defined. Here we show that SUMO-conjugating enzyme Ubc9 interacts with a conserved GKAE motif in ROR-γt to induce SUMOylation of ROR-γt and suppress IL-17 expression. Th17 cells expressing SUMOylation-defective ROR-γt are highly colitogenic upon transfer to Rag1–/– mice. Mechanistically, SUMOylation of ROR-γt facilitates the binding of HDAC2 to the IL-17 promoter and represses IL-17 transcription. Mice with conditional deletion of HDAC2 in CD4+ T cells have elevated IL-17 expression and severe colitis. The identification of the Ubc9/ROR-γt/HDAC2 axis that governs IL-17 expression may open new venues for the development of therapeutic measures for inflammatory disorders., Interleukin-17 (IL-17)-secreting CD4 T cells (Th17) are induced by the master transcription factor RORγt, and are important for anti-fungal immunity and inflammatory responses. Here the authors show that Ubc9-mediated SUMOylation of RORγt induces HDAC2 binding to IL-17 promoter for suppressing IL-17 production in Th17 cells.

SMAD4 constrains progression of Pten-null prostate cancer and serves as a common downstream node of transforming growth factor β (TGFβ) and bone morphogenetic protein (BMP) pathways. Here, we dissected the roles of TGFβ receptor II (TGFBR2) and BMP receptor II (BMPR2) using a Pten-null prostate cancer model. These studies demonstrated that the molecular actions of TGFBR2 result in both SMAD4-dependent constraint of proliferation and SMAD4-independent activation of apoptosis. In contrast, BMPR2 deletion extended survival relative to Pten deletion alone, establishing its promoting role in BMP6-driven prostate cancer progression. These analyses reveal the complexity of TGFβ–BMP signaling and illuminate potential therapeutic targets for prostate cancer.

Genetic inactivation of PTEN is common in prostate cancer and correlates with poorer prognosis. We previously identified CHD1 as an essential gene in PTEN-deficient cancer cells. Here, we sought definitive in vivo genetic evidence for, and mechanistic understanding of, the essential role of CHD1 in PTEN-deficient prostate cancer. In Pten and Pten/Smad4 genetically engineered mouse models, prostate-specific deletion of Chd1 resulted in markedly delayed tumor progression and prolonged survival. Chd1 deletion was associated with profound tumor microenvironment (TME) remodeling characterized by reduced myeloid-derived suppressor cells (MDSC) and increased CD8+ T cells. Further analysis identified IL6 as a key transcriptional target of CHD1, which plays a major role in recruitment of immunosuppressive MDSCs. Given the prominent role of MDSCs in suppressing responsiveness to immune checkpoint inhibitors (ICI), our genetic and tumor biological findings support combined testing of anti-IL6 and ICI therapies, specifically in PTEN-deficient prostate cancer. Significance: We demonstrate a critical role of CHD1 in MDSC recruitment and discover CHD1/IL6 as a major regulator of the immunosuppressive TME of PTEN-deficient prostate cancer. Pharmacologic inhibition of IL6 in combination with immune checkpoint blockade elicits robust antitumor responses in prostate cancer. This article is highlighted in the In This Issue feature, p. 1241

Penile squamous cell carcinoma (PSCC) accounts for over 95% of penile malignancies and causes significant mortality and morbidity in developing countries. Molecular mechanisms and therapies of PSCC are understudied, owing to scarcity of laboratory models. Herein, we describe a genetically engineered mouse model of PSCC, by co-deletion of Smad4 and Apc in the androgen-responsive epithelium of the penis. Mouse PSCC fosters an immunosuppressive microenvironment with myeloid-derived suppressor cells (MDSCs) as a dominant population. Preclinical trials in the model demonstrate synergistic efficacy of immune checkpoint blockade with the MDSC-diminishing drugs cabozantinib or celecoxib. A critical clinical problem of PSCC is chemoresistance to cisplatin, which is induced by Pten deficiency on the backdrop of Smad4/Apc co-deletion. Drug screen studies informed by targeted proteomics identify a few potential therapeutic strategies for PSCC. Our studies have established what we believe to be essential resources for studying PSCC biology and developing therapeutic strategies., Penile squamous cell carcinoma (PSCC) is a cancer that is associated with significant mortality. Here, the authors develop a mouse model of PSCC by co-deletion of Smad4 and Apc in the androgen-responsive penile epithelium, and show synergistic efficacy of checkpoint therapy with cabozantinib or celecoxib in their model.

A hallmark of pancreatic ductal adenocarcinoma (PDAC) is an exuberant stroma comprised of diverse cell types that enable or suppress tumor progression. Here, we explored the role of oncogenic KRAS in protumorigenic signaling interactions between cancer cells and host cells. We show that KRAS mutation (KRAS*) drives cell-autonomous expression of type I cytokine receptor complexes (IL2rγ–IL4rα and IL2rγ–IL13rα1) in cancer cells that in turn are capable of receiving cytokine growth signals (IL4 or IL13) provided by invading Th2 cells in the microenvironment. Early neoplastic lesions show close proximity of cancer cells harboring KRAS* and Th2 cells producing IL4 and IL13. Activated IL2rγ–IL4rα and IL2rγ–IL13rα1 receptors signal primarily via JAK1–STAT6. Integrated transcriptomic, chromatin occupancy, and metabolomic studies identified MYC as a direct target of activated STAT6 and that MYC drives glycolysis. Thus, paracrine signaling in the tumor microenvironment plays a key role in the KRAS*-driven metabolic reprogramming of PDAC. Significance: Type II cytokines, secreted by Th2 cells in the tumor microenvironment, can stimulate cancer cell–intrinsic MYC transcriptional upregulation to drive glycolysis. This KRAS*-driven heterotypic signaling circuit in the early and advanced tumor microenvironment enables cooperative protumorigenic interactions, providing candidate therapeutic targets in the KRAS* pathway for this intractable disease.

Mitochondrial dysfunction is behind several neurodegenerative diseases, including Alzheimer disease (AD). Accumulation of damaged mitochondria is already observed at the early stages of AD and has been linked to impaired mitophagy, but the mechanisms underlying this alteration are still not fully known. In our recent study, we show that intracellular cholesterol enrichment can downregulate amyloid beta (Aβ)-induced mitophagy. Mitochondrial glutathione depletion resulting from high cholesterol levels promotes PINK1 (PTEN induced kinase 1)-mediated mitophagosome formation; however, mitophagy flux is ultimately disrupted, most likely due to fusion deficiency of endosomes-lysosomes caused by cholesterol. Meanwhile, in APP-PSEN1-SREBF2 mice, an AD mouse model that overexpresses the cholesterol-related transcription factor SREBF2, cholesterol accumulation prompts an oxidative- and age-dependent cytosolic aggregation of the mitophagy adaptor OPTN (optineurin), which prevents mitophagosome formation despite enhanced PINK1-PRKN/parkin signaling. Hippocampal neurons from postmortem brain of AD individuals reproduce the progressive accumulation of OPTN in aggresome-like structures accompanied by high levels of mitochondrial cholesterol in advanced stages of the disease. Overall, these data provide new insights into the impairment of the PINK1-PRKN mitophagy pathway in AD and suggest the combination of mitophagy inducers with strategies focused on restoring the cholesterol homeostasis and mitochondrial redox balance as a potential disease-modifying therapy for AD.

The signaling mechanisms between prostate cancer cells and infiltrating immune cells may illuminate novel therapeutic approaches. Here, utilizing a prostate adenocarcinoma model driven by loss of Pten and Smad4, we identify polymorphonuclear myeloid-derived suppressor cells (MDSC) as the major infiltrating immune cell type, and depletion of MDSCs blocks progression. Employing a novel dual reporter prostate cancer model, epithelial and stromal transcriptomic profiling identified CXCL5 as a cancer-secreted chemokine to attract CXCR2-expressing MDSCs, and, correspondingly, pharmacologic inhibition of CXCR2 impeded tumor progression. Integrated analyses identified hyperactivated Hippo–YAP signaling in driving CXCL5 upregulation in cancer cells through the YAP–TEAD complex and promoting MDSC recruitment. Clinicopathologic studies reveal upregulation and activation of YAP1 in a subset of human prostate tumors, and the YAP1 signature is enriched in primary prostate tumor samples with stronger expression of MDSC-relevant genes. Together, YAP-driven MDSC recruitment via heterotypic CXCL5–CXCR2 signaling reveals an effective therapeutic strategy for advanced prostate cancer. Significance: We demonstrate a critical role of MDSCs in prostate tumor progression and discover a cancer cell nonautonomous function of the Hippo–YAP pathway in regulation of CXCL5, a ligand for CXCR2-expressing MDSCs. Pharmacologic elimination of MDSCs or blocking the heterotypic CXCL5–CXCR2 signaling circuit elicits robust antitumor responses and prolongs survival. Cancer Discov; 6(1); 80–95. ©2015 AACR. This article is highlighted in the In This Issue feature, p. 1

Purpose While leptin has been associated with various psycho-physiological functions, the molecular network in leptin-mediated mood regulation remains elusive. Methods Anxiolytic behaviors and tyrosine hydroxylase (TH) levels were examined after leptin administration. Functional roles of STAT3 and FoxO1 in regulation of TH expression were investigated using in vivo and in vitro systems. A series of animal behavioral tests using dopaminergic neuron-specific FoxO1 KO (FoxO1 KODAT) were performed and investigated the roles of FoxO1 in regulation of mood behaviors. Results Here, we show that administration of leptin induces anxiolytic-like phenotype through the activation of signal transducer and activator of transcription 3 (STAT3) and the inhibition of forkhead box protein O1 (FoxO1) in dopaminergic (DA) neurons of the midbrain. Specifically, STAT3 and FoxO1 directly bind to and exert opposing effects on tyrosine hydroxylase (TH) expression, where STAT3 acts as an enhancer and FoxO1 acts as a prominent repressor. Accordingly, suppression of the prominent suppressor FoxO1 by leptin strongly increased TH expression. Furthermore, our previous results showed that specific deletion of FoxO1 in DA neurons (FoxO1 KODAT) led to a profound elevation of TH activity and dopamine contents. Finally, FoxO1 KODAT mice exhibited enhanced leptin sensitivity as well as displayed reduced anxiety- and depression-like behaviors. Conclusions This work establishes a novel molecular mechanism of mood behavior regulation by leptin and suggests FoxO1 suppression by leptin might be a key for leptin-induced behavioral manifestation in DA neurons.

Despite the high long-term survival in localized prostate cancer, metastatic prostate cancer remains largely incurable even after intensive multimodal therapy. The lethality of advanced disease is driven by the lack of therapeutic regimens capable of generating durable responses in the setting of extreme tumor heterogeneity on the genetic and cell biological levels. Here, we review available prostate cancer model systems, the prostate cancer genome atlas, cellular and functional heterogeneity in the tumor microenvironment, tumor-intrinsic and tumor-extrinsic mechanisms underlying therapeutic resistance, and technological advances focused on disease detection and management. These advances, along with an improved understanding of the adaptive responses to conventional cancer therapies, anti-androgen therapy, and immunotherapy, are catalyzing development of more effective therapeutic strategies for advanced disease. In particular, knowledge of the heterotypic interactions between and coevolution of cancer and host cells in the tumor microenvironment has illuminated novel therapeutic combinations with a strong potential for more durable therapeutic responses and eventual cures for advanced disease. Improved disease management will also benefit from artificial intelligence-based expert decision support systems for proper standard of care, prognostic determinant biomarkers to minimize overtreatment of localized disease, and new standards of care accelerated by next-generation adaptive clinical trials.

Clustered regularly interspaced short palindromic repeats (CRISPR), CRISPR interference and programmable base editing have transformed the manipulation of eukaryotic genomes for potential therapeutic applications

Advanced prostate cancer displays conspicuous chromosomal instability and rampant copy number aberrations, yet the identity of functional drivers resident in many amplicons remain elusive. Here, we implemented a functional genomics approach to identify new oncogenes involved in prostate cancer progression. Through integrated analyses of focal amplicons in large prostate cancer genomic and transcriptomic datasets as well as genes upregulated in metastasis, 276 putative oncogenes were enlisted into an in vivo gain-of-function tumorigenesis screen. Among the top positive hits, we conducted an in-depth functional analysis on Pygopus family PHD finger 2 (PYGO2), located in the amplicon at 1q21.3. PYGO2 overexpression enhances primary tumor growth and local invasion to draining lymph nodes. Conversely, PYGO2 depletion inhibits prostate cancer cell invasion in vitro and progression of primary tumor and metastasis in vivo. In clinical samples, PYGO2 upregulation associated with higher Gleason score and metastasis to lymph nodes and bone. Silencing PYGO2 expression in patient-derived xenograft models impairs tumor progression. Finally, PYGO2 is necessary to enhance the transcriptional activation in response to ligand-induced Wnt/β-catenin signaling. Together, our results indicate that PYGO2 functions as a driver oncogene in the 1q21.3 amplicon and may serve as a potential prognostic biomarker and therapeutic target for metastatic prostate cancer. Significance: Amplification/overexpression of PYGO2 may serve as a biomarker for prostate cancer progression and metastasis. Cancer Res; 78(14); 3823–33. ©2018 AACR.

Purpose: Prostate cancers show remarkable resistance to emerging immunotherapies, partly due to tolerogenic STAT3 signaling in tumor-associated myeloid cells. Here, we describe a novel strategy combining STAT3 inhibition with Toll-like Receptor 9 (TLR9) stimulation to unleash immune response against prostate cancers regardless of the genetic background. Experimental Design: We developed and validated a conjugate of the STAT3 antisense oligonucleotide (ASO) tethered to immunostimulatory TLR9 agonist (CpG oligonucleotide) to improve targeting of human and mouse prostate cancer and myeloid immune cells, such as myeloid-derived suppressor cells (MDSC). Results: CpG-STAT3ASO conjugates showed improved biodistribution and potency of STAT3 knockdown in target cells in vitro and in vivo. Systemic administration of CpG-STAT3ASO (5 mg/kg) eradicated bone-localized, Ras/Myc-driven, and Ptenpc−/−Smad4pc−/−Trp53c−/− prostate tumors in the majority of treated mice. These antitumor effects were primarily immune-mediated and correlated with an increased ratio of CD8+ to regulatory T cells and reduced pSTAT3+/PD-L1+ MDSCs. Both innate and adaptive immunity contributed to systemic antitumor responses as verified by the depletion of Gr1+ myeloid cells and CD8+ and CD4+ T cells, respectively. Importantly, only the bifunctional CpG-STAT3ASO, but not control CpG oligonucleotides, STAT3ASO alone, or the coinjection of both oligonucleotides, succeeded in recruiting neutrophils and CD8+ T cells into tumors. Thus, the concurrence of TLR9 activation with STAT3 inhibition in the same cellular compartment is indispensable for overcoming tumor immune tolerance and effective antitumor immunity against prostate cancer. Conclusions: The bifunctional, immunostimulatory, and tolerance-breaking design of CpG-STAT3ASO offers a blueprint for the development of effective and safer oligonucleotide strategies for treatment of immunologically “cold” human cancers.

Metabolic reprograming is an emerging hallmark of tumor biology and an actively pursued opportunity in discovery of oncology drugs. Extensive efforts have focused on therapeutic targeting of glycolysis, whereas drugging mitochondrial oxidative phosphorylation (OXPHOS) has remained largely unexplored, partly owing to an incomplete understanding of tumor contexts in which OXPHOS is essential. Here, we report the discovery of IACS-010759, a clinical-grade small-molecule inhibitor of complex I of the mitochondrial electron transport chain. Treatment with IACS-010759 robustly inhibited proliferation and induced apoptosis in models of brain cancer and acute myeloid leukemia (AML) reliant on OXPHOS, likely owing to a combination of energy depletion and reduced aspartate production that leads to impaired nucleotide biosynthesis. In models of brain cancer and AML, tumor growth was potently inhibited in vivo following IACS-010759 treatment at well-tolerated doses. IACS-010759 is currently being evaluated in phase 1 clinical trials in relapsed/refractory AML and solid tumors.

Blood-based biomarkers for early detection of colorectal cancer could complement current approaches to colorectal cancer screening. We previously identified the APC-binding protein MAPRE1 as a potential colorectal cancer biomarker. Here, we undertook a case–control validation study to determine the performance of MAPRE1 in detecting early colorectal cancer and colon adenoma and to assess the potential relevance of additional biomarker candidates. We analyzed plasma samples from 60 patients with adenomas, 30 with early colorectal cancer, 30 with advanced colorectal cancer, and 60 healthy controls. MAPRE1 and a set of 21 proteins with potential biomarker utility were assayed using high-density antibody arrays, and carcinoembryonic antigen (CEA) was assayed using ELISA. The biologic significance of the candidate biomarkers was also assessed in colorectal cancer mouse models. Plasma MAPRE1 levels were significantly elevated in both patients with adenomas and patients with colorectal cancer compared with controls (P < 0.0001). MAPRE1 and CEA together yielded an area under the curve of 0.793 and a sensitivity of 0.400 at 95% specificity for differentiating early colorectal cancer from controls. Three other biomarkers (AK1, CLIC1, and SOD1) were significantly increased in both adenoma and early colorectal cancer patient plasma samples and in plasma from colorectal cancer mouse models at preclinical stages compared with controls. The combination of MAPRE1, CEA, and AK1 yielded sensitivities of 0.483 and 0.533 at 90% specificity and sensitivities of 0.350 and 0.467 at 95% specificity for differentiating adenoma and early colorectal cancer, respectively, from healthy controls. These findings suggest that MAPRE1 can contribute to the detection of early-stage colorectal cancer and adenomas together with other biomarkers. Cancer Prev Res; 8(11); 1112–9. ©2015 AACR.

SummaryMyelodysplastic syndrome (MDS) risk correlates with advancing age, therapy-induced DNA damage, and/or shorter telomeres, but whether telomere erosion directly induces MDS is unknown. Here, we provide the genetic evidence that telomere dysfunction-induced DNA damage drives classical MDS phenotypes and alters common myeloid progenitor (CMP) differentiation by repressing the expression of mRNA splicing/processing genes, including SRSF2. RNA-seq analyses of telomere dysfunctional CMP identified aberrantly spliced transcripts linked to pathways relevant to MDS pathogenesis such as genome stability, DNA repair, chromatin remodeling, and histone modification, which are also enriched in mouse CMP haploinsufficient for SRSF2 and in CD34+ CMML patient cells harboring SRSF2 mutation. Together, our studies establish an intimate link across telomere biology, aberrant RNA splicing, and myeloid progenitor differentiation.

Autophagy is a conserved catabolic pathway with emerging functions in mammalian neurodevelopment and human neurodevelopmental diseases. The mechanisms controlling autophagy in neuronal development are not fully understood. Here, we found that conditional deletion of the Forkhead Box O transcription factors FoxO1, FoxO3, and FoxO4 strongly impaired autophagic flux in developing neurons of the adult mouse hippocampus. Moreover, FoxO deficiency led to altered dendritic morphology, increased spine density, and aberrant spine positioning in adult-generated neurons. Strikingly, pharmacological induction of autophagy was sufficient to correct abnormal dendrite and spine development of FoxO-deficient neurons. Collectively, these findings reveal a novel link between FoxO transcription factors, autophagic flux, and maturation of developing neurons.

An integrated genomic and functional analysis to elucidate DNA damage signaling factors promoting self-renewal of glioma stem cells (GSCs) identified proliferating cell nuclear antigen (PCNA)-associated factor (PAF) up-regulation in glioblastoma. PAF is preferentially overexpressed in GSCs. Its depletion impairs maintenance of self-renewal without promoting differentiation and reduces tumor-initiating cell frequency. Combined transcriptomic and metabolomic analyses revealed that PAF supports GSC maintenance, in part, by influencing DNA replication and pyrimidine metabolism pathways. PAF interacts with PCNA and regulates PCNA-associated DNA translesion synthesis (TLS); consequently, PAF depletion in combination with radiation generated fewer tumorspheres compared with radiation alone. Correspondingly, pharmacological impairment of DNA replication and TLS phenocopied the effect of PAF depletion in compromising GSC self-renewal and radioresistance, providing preclinical proof of principle that combined TLS inhibition and radiation therapy may be a viable therapeutic option in the treatment of glioblastoma multiforme (GBM).

E26 transformation-specific transcription factor ERG is aberrantly overexpressed in approximately 50% of all human prostate cancer due to TMPRSS2-ERG gene rearrangements. However, mice with prostate-specific transgenic expression of prostate cancer–associated ERG alone fail to develop prostate cancer, highlighting that ERG requires other lesions to drive prostate tumorigenesis. Forkhead box (FOXO) transcription factor FOXO1 is a tumor suppressor that is frequently inactivated in human prostate cancer. Here, we demonstrate that FOXO1, but not other FOXO proteins (FOXO3 and FOXO4), binds and inhibits the transcriptional activity of prostate cancer–associated ERG independently of FOXO1 transcriptional activity. Knockdown of endogenous FOXO1 increased invasion of TMPRSS2–ERG fusion–positive VCaP cells, an effect completely abolished by ERG knockdown. Patient specimen analysis demonstrated that FOXO1 and ERG protein expression inversely correlated in a subset of human prostate cancer. Although human ERG transgene expression or homozygous deletion of Foxo1 alone in the mouse prostate failed to promote tumorigenesis, concomitant ERG transgene expression and Foxo1 deletion resulted in upregulation of ERG target genes, increased cell proliferation, and formation of high-grade prostatic intraepithelial neoplasia. Overall, we provide biochemical and genetic evidence that aberrantly activated ERG cooperates with FOXO1 deficiency to promote prostate tumorigenesis and cell invasion. Our findings enhance understanding of prostate cancer etiology and suggest that the FOXO1–ERG signaling axis can be a potential target for treatment of prostate cancer. Cancer Res; 77(23); 6524–37. ©2017 AACR.

Kras activation and p16 inactivation are required to develop pancreatic ductal adenocarcinoma (PDAC). However, the biochemical mechanisms underlying these double alterations remain unclear. Here we discover that NAD(P)H oxidase 4 (NOX4), an enzyme known to catalyse the oxidation of NAD(P)H, is upregulated when p16 is inactivated by looking at gene expression profiling studies. Activation of NOX4 requires catalytic subunit p22phox, which is upregulated following Kras activation. Both alterations are also detectable in PDAC cell lines and patient specimens. Furthermore, we show that elevated NOX4 activity accelerates oxidation of NADH and supports increased glycolysis by generating NAD+, a substrate for GAPDH-mediated glycolytic reaction, promoting PDAC cell growth. Mechanistically, NOX4 was induced through p16-Rb-regulated E2F and p22phox was induced by KrasG12V-activated NF-κB. In conclusion, we provide a biochemical explanation for the cooperation between p16 inactivation and Kras activation in PDAC development and suggest that NOX4 is a potential therapeutic target for PDAC., Kras activation and p16 inactivation cooperatively promote pancreatic cancer progression. Here, the authors show that such cooperation depends upon an increased expression of the NAD(P)H oxidase NOX4 achieved through transcription factors independently regulated by the two oncogenic genetic alterations.

A key feature of high-grade serous ovarian carcinoma (HGSOC) is frequent amplification of the 3q26 locus harboring PRKC-ι (PRKCI). Here, we show that PRKCI is also expressed in early fallopian tube lesions, called serous tubal intraepithelial carcinoma. Transgenic mouse studies establish PRKCI as an ovarian cancer-specific oncogene. Mechanistically, we show that the oncogenic activity of PRKCI relates in part to the up-regulation of TNFα to promote an immune-suppressive tumor microenvironment characterized by an abundance of myeloid-derived suppressor cells and inhibition of cytotoxic T-cell infiltration. Furthermore, system-level and functional analyses identify YAP1 as a downstream effector in tumor progression. In human ovarian cancers, high PRKCI expression also correlates with high expression of TNFα and YAP1 and low infiltration of cytotoxic T cells. The PRKCI–YAP1 regulation of the tumor immunity provides a therapeutic strategy for highly lethal ovarian cancer.

Malignant neoplasms evolve in response to changes in oncogenic signalling(1). Cancer cell plasticity in response to evolutionary pressures is fundamental to tumour progression and the development of therapeutic resistance(2,3). Here we elucidate the molecular and cellular mechanisms of cancer cell plasticity in a conditional oncogenic Kras mouse model of pancreatic ductal adenocarcinoma (PDAC), a malignancy that displays considerable phenotypic diversity and morphological heterogeneity. In this model, stochastic extinction of oncogenic Kras signalling and emergence of Kras-independent escaper populations (cells that acquire oncogenic properties) are associated with de-differentiation and aggressive biological behaviour. Transcriptomic and functional analyses of Kras-independent escapers reveal the presence of Smarcb1–Myc-network-driven mesenchymal reprogramming and independence from MAPK signalling. A somatic mosaic model of PDAC, which allows time-restricted perturbation of cell fate, shows that depletion of Smarcb1 activates the Myc network, driving an anabolic switch that increases protein metabolism and adaptive activation of endoplasmic-reticulum-stress-induced survival pathways. Elevated protein turnover renders mesenchymal sub-populations highly susceptible to pharmacological and genetic perturbation of the cellular proteostatic machinery and the IRE1-α–MKK4 arm of the endoplasmic-reticulum-stress-response pathway. Specifically, combination regimens that impair the unfolded protein responses block the emergence of aggressive mesenchymal subpopulations in mouse and patient-derived PDAC models. These molecular and biological insights inform a potential therapeutic strategy for targeting aggressive mesenchymal features of PDAC.

Activating mutations in KRAS are among the most frequent events in diverse human carcinomas and are particularly prominent in human pancreatic ductal adenocarcinoma (PDAC). An inducible Kras(G12D)-driven mouse model of PDAC has established a critical role for sustained Kras(G12D) expression in tumor maintenance, providing a model to determine the potential for and the underlying mechanisms of Kras(G12D)-independent PDAC recurrence. Here, we show that some tumors undergo spontaneous relapse and are devoid of Kras(G12D) expression and downstream canonical MAPK signaling and instead acquire amplification and overexpression of the transcriptional coactivator Yap1. Functional studies established the role of Yap1 and the transcriptional factor Tead2 in driving Kras(G12D)-independent tumor maintenance. The Yap1/Tead2 complex acts cooperatively with E2F transcription factors to activate a cell cycle and DNA replication program. Our studies, along with corroborating evidence from human PDAC models, portend a novel mechanism of escape from oncogenic Kras addiction in PDAC.

Heterotypic interactions across diverse cell types can enable tumor progression and hold the potential to expand therapeutic interventions. Here, combined profiling and functional studies of glioma cells in glioblastoma multiforme (GBM) models establish that PTEN deficiency activates YAP1, which directly upregulates lysyl oxidase (LOX) expression. Mechanistically, secreted LOX functions as a potent macrophage chemoattractant via activation of the β1 integrin-PYK2 pathway in macrophages. These infiltrating macrophages secrete SPP1, which sustains glioma cell survival and stimulates angiogenesis. In PTEN-null GBM models, LOX inhibition markedly suppresses macrophage infiltration and tumor progression. Correspondingly, YAP1-LOX and β1 integrin-SPP1 signaling correlates positively with higher macrophage density and lower overall survival in GBM patients. This symbiotic glioma-macrophage interplay provides therapeutic targets specifically for PTEN-deficient GBM.

The biological functions and mechanisms of oncogenic KRASG12D (KRAS∗) in resistance to immune checkpoint blockade (ICB) therapy are not fully understood. We demonstrate that KRAS∗ represses the expression of interferon regulatory factor 2 (IRF2), which in turn directly represses CXCL3 expression. KRAS∗-mediated repression of IRF2 results in high expression of CXCL3, which binds to CXCR2 on myeloid-derived suppressor cells and promotes their migration to the tumor microenvironment. Anti-PD-1 resistance of KRAS∗-expressing tumors can be overcome by enforced IRF2 expression or by inhibition of CXCR2. Colorectal cancer (CRC) showing higher IRF2 expression exhibited increased responsiveness to anti-PD-1 therapy. The KRAS∗-IRF2-CXCL3-CXCR2 axis provides a framework for patient selection and combination therapies to enhance the effectiveness of ICB therapy in CRC.

Purpose: gp96 (grp94) is a key downstream chaperone in the endoplasmic reticulum (ER) to mediate unfolded protein response (UPR) and the pathogenesis of multiple myeloma is closely linked to dysregulated UPR. In this study, we aimed to determine the roles of gp96 in the initiation and progression of multiple myeloma in vivo and in vitro. Experimental Design: We generated a mouse model with overexpression of XBP1s and conditional deletion of gp96 in B-cell compartment simultaneously to identify the roles of gp96 in the development of multiple myeloma in vivo. Using a short hairpin RNA (shRNA) system, we silenced gp96 in multiple human multiple myeloma cells and examined the effect of gp96 knockdown on multiple myeloma cells by cell proliferation, cell-cycle analysis, apoptosis assay, immunohistochemistry, and human myeloma xenograft model. The anticancer activity of gp96 selective inhibitor, WS13, was evaluated by apoptosis assay and MTT assay. Results: Genetic deletion of gp96 in XBP1s-Tg mice attenuates multiple myeloma. Silencing of gp96 causes severe compromise in human multiple myeloma cell growth through inhibiting Wnt-LRP-survivin pathway. We also confirmed that knockdown of gp96 decreased human multiple myeloma growth in a murine xenograft model. The targeted gp96 inhibitor induced apoptosis and blocked multiple myeloma cell growth, but did not induce apoptosis in pre-B leukemic cells. We have demonstrated that myeloma growth is dependent on gp96 both genetically and pharmacologically. Conclusions: gp96 is essential for multiple myeloma cell proliferation and survival, suggesting that gp96 is a novel therapeutic target for multiple myeloma. Clin Cancer Res; 19(22); 6242–51. ©2013 AACR.

The pyruvate kinase M2 isoform (PKM2) is expressed in cancer and plays a role in regulating anabolic metabolism. To determine whether PKM2 is required for tumor formation or growth, we generated mice with a conditional allele that abolishes PKM2 expression without disrupting PKM1 expression. PKM2 deletion accelerated mammary tumor formation in a Brca1-loss-driven model of breast cancer. PKM2 null tumors displayed heterogeneous PKM1 expression, with PKM1 found in nonproliferating tumor cells and no detectable pyruvate kinase expression in proliferating cells. This suggests that PKM2 is not necessary for tumor cell proliferation and implies that the inactive state of PKM2 is associated with the proliferating cell population within tumors, whereas nonproliferating tumor cells require active pyruvate kinase. Consistent with these findings, variable PKM2 expression and heterozygous PKM2 mutations are found in human tumors. These data suggest that regulation of PKM2 activity supports the different metabolic requirements of proliferating and nonproliferating tumor cells., National Institutes of Health (U.S.) (Grant R01CA168653), National Institutes of Health (U.S.) (Grant 5P01CA117969), National Institutes of Health (U.S.) (Grant P30CA147882), National Institutes of Health (U.S.) (Grant 5P30CA14051), National Institutes of Health (U.S.) (Grant 5K08CA136983), National Institutes of Health (U.S.) (Grant DK059635), National Institutes of Health (U.S.) (Grant R01DK092606), National Institutes of Health (U.S.) (Grant R00CA131472), National Institutes of Health (U.S.) (Grant R01GM056203), American Diabetes Association (Grant 7-12-BS-09), Smith Family Foundation, Burroughs Wellcome Fund, Damon Runyon Cancer Research Foundation, Stern Family

Background/Aims Impaired regulation of apoptosis has been suggested to play a role in the pathogenesis of Primary Biliary Cirrhosis (PBC). In this study, we analysed a signalling pathway that comprises the transcription factor FoxO3a and its downstream target Bim, a Bcl-2 interacting mediator of apoptosis. Materials & Methods The tissues examined included livers explanted from patients with cirrhotic PBC, primary sclerosing cholangitis (PSC), alcoholic liver disease (ALD) and liver biopsies from patients with non-cirrhotic PBC. Large margin resections of hepatocellular carcinoma were used as controls. Results Expression of FoxO3a and Bim mRNA was significantly enhanced in both non-cirrhotic and end-stage PBC (2.2-fold and 4.3-fold increases, respectively), but not in the other disorders. Similarly, FOXO3a protein level was increased in end-stage PBC (P

A significant fraction of patients with advanced prostate cancer treated with androgen deprivation therapy experience relapse with relentless progression to lethal metastatic castration-resistant prostate cancer (mCRPC). Immune checkpoint blockade using antibodies against cytotoxic-T-lymphocyte-associated protein 4 (CTLA4) or programmed cell death 1/programmed cell death 1 ligand 1 (PD1/PD-L1) generates durable therapeutic responses in a significant subset of patients across a variety of cancer types. However, mCRPC showed overwhelming de novo resistance to immune checkpoint blockade, motivating a search for targeted therapies that overcome this resistance. Myeloid-derived suppressor cells (MDSCs) are known to play important roles in tumour immune evasion. The abundance of circulating MDSCs correlates with prostate-specific antigen levels and metastasis in patients with prostate cancer. Mouse models of prostate cancer show that MDSCs (CD11b+Gr1+) promote tumour initiation and progression. These observations prompted us to hypothesize that robust immunotherapy responses in mCRPC may be elicited by the combined actions of immune checkpoint blockade agents together with targeted agents that neutralize MDSCs yet preserve T-cell function. Here we develop a novel chimaeric mouse model of mCRPC to efficiently test combination therapies in an autochthonous setting. Combination of anti-CTLA4 and anti-PD1 engendered only modest efficacy. Targeted therapy against mCRPC-infiltrating MDSCs, using multikinase inhibitors such as cabozantinib and BEZ235, also showed minimal anti-tumour activities. Strikingly, primary and metastatic CRPC showed robust synergistic responses when immune checkpoint blockade was combined with MDSC-targeted therapy. Mechanistically, combination therapy efficacy stemmed from the upregulation of interleukin-1 receptor antagonist and suppression of MDSC-promoting cytokines secreted by prostate cancer cells. These observations illuminate a clinical path hypothesis for combining immune checkpoint blockade with MDSC-targeted therapies in the treatment of mCRPC.

Here, Boutin et al. investigated the role of oncogenic KRAS in colorectal cancer (CRC). They generated a mouse model of metastatic CRC that harbors an inducible oncogenic Kras allele (Krasmut) and conditional null alleles of Apc and Trp53 (iKAP), which provides genetic evidence that Krasmut drives CRC invasion and maintenance of metastases.

The genome of pancreatic ductal adenocarcinoma (PDAC) frequently contains deletions of tumour suppressor gene loci, most notably SMAD4, which is homozygously deleted in nearly one-third of cases. As loss of neighbouring housekeeping genes can confer collateral lethality, we sought to determine whether loss of the metabolic gene malic enzyme 2 (ME2) in the SMAD4 locus would create cancer-specific metabolic vulnerability upon targeting of its paralogous isoform ME3. The mitochondrial malic enzymes (ME2 and ME3) are oxidative decarboxylases that catalyse the conversion of malate to pyruvate and are essential for NADPH regeneration and reactive oxygen species homeostasis. Here we show that ME3 depletion selectively kills ME2-null PDAC cells in a manner consistent with an essential function for ME3 in ME2-null cancer cells. Mechanistically, integrated metabolomic and molecular investigation of cells deficient in mitochondrial malic enzymes revealed diminished NADPH production and consequent high levels of reactive oxygen species. These changes activate AMP activated protein kinase (AMPK), which in turn directly suppresses sterol regulatory element-binding protein 1 (SREBP1)-directed transcription of its direct targets including the BCAT2 branched-chain amino acid transaminase 2) gene. BCAT2 catalyses the transfer of the amino group from branched-chain amino acids to α-ketoglutarate (α-KG) thereby regenerating glutamate, which functions in part to support de novo nucleotide synthesis. Thus, mitochondrial malic enzyme deficiency, which results in impaired NADPH production, provides a prime 'collateral lethality' therapeutic strategy for the treatment of a substantial fraction of patients diagnosed with this intractable disease.

Autophagy plays a critical role in cell survival from prolonged starvation and recycling of aggregated proteins and damaged organelles. One of the essential genes involved in the autophagic initiation is autophagy-related 14 (Atg14), also called Barkor for Beclin 1-associated autophagy-related key regulator. Although its crucial role in the autophagic process has been reported, the gene regulation of Atg14 and its metabolic functions remain unclear. In this work we have identified that the Atg14 gene is regulated by forkhead box O (FoxO) transcription factors and circadian rhythms in the mouse liver. Luciferase reporter analyses and chromatin immunoprecipitation assays have revealed well conserved cis-elements for FoxOs and Clock/Bmal1 in the proximal promoter of the Atg14 gene. To examine the functions of hepatic Atg14, we have performed the gene knockdown and overexpression in the mouse livers. Remarkably, knockdown of Atg14 leads to elevated levels of triglycerides in the liver and serum as well. Conversely, overexpression of Atg14 improves hypertriglyceridemia in both high fat diet-treated wild-type mice and FoxO1/3/4 liver-specific knock-out mice. In summary, our data suggest that Atg14 is a new target gene of FoxOs and the core clock machinery, and this gene plays an important role in hepatic lipid metabolism.

Glioblastoma multiforme (GBM) is the most aggressive of the astrocytic malignancies and the most common intracranial tumor in adults. Although the epidermal growth factor receptor (EGFR) is overexpressed and/or mutated in at least 50% of GBM cases and is required for tumor maintenance in animal models, EGFR inhibitors have thus far failed to deliver significant responses in GBM patients. One inherent resistance mechanism in GBM is the coactivation of multiple receptor tyrosine kinases, which generates redundancy in activation of phosphoinositide-3′-kinase (PI3K) signaling. Here we demonstrate that the phosphatase and tensin homolog deleted on chromosome 10 (PTEN) tumor suppressor is frequently phosphorylated at a conserved tyrosine residue, Y240, in GBM clinical samples. Phosphorylation of Y240 is associated with shortened overall survival and resistance to EGFR inhibitor therapy in GBM patients and plays an active role in mediating resistance to EGFR inhibition in vitro. Y240 phosphorylation can be mediated by both fibroblast growth factor receptors and SRC family kinases (SFKs) but does not affect the ability of PTEN to antagonize PI3K signaling. These findings show that, in addition to genetic loss and mutation of PTEN, its modulation by tyrosine phosphorylation has important implications for the development and treatment of GBM.

The transcription factor FOXO1 plays a central role in metabolic homeostasis by regulating leptin and insulin activity in many cell types, including neurons. However, the neurons mediating these effects and the identity of the molecular targets through which FOXO1 regulates metabolism remain to be defined. Here, we show that the ventral medial nucleus of the hypothalamus (VMH) is a key site of FOXO1 action. We found that mice lacking FOXO1 in steroidogenic factor 1 (SF-1) neurons of the VMH are lean due to increased energy expenditure. The mice also failed to appropriately suppress energy expenditure in response to fasting. Furthermore, these mice displayed improved glucose tolerance due to increased insulin sensitivity in skeletal muscle and heart. Gene expression profiling and sequence analysis revealed several pathways regulated by FOXO1. In addition, we identified the nuclear receptor SF-1 as a direct FOXO1 transcriptional target in the VMH. Collectively, our data suggest that the transcriptional networks modulated by FOXO1 in VMH neurons are key components in the regulation of energy balance and glucose homeostasis.

Progressive DNA damage and mitochondrial decline are both considered to be prime instigators of natural ageing. Traditionally, these two pathways have been viewed largely in isolation. However, recent studies have revealed a molecular circuit that directly links DNA damage to compromised mitochondrial biogenesis and function via p53. This axis of ageing may account for both organ decline and disease development associated with advanced age and could illuminate a path for the development of relevant therapeutics.

Restoration of regulated insulin secretion is the ultimate goal of therapy for type 1 diabetes. Here, we show that, unexpectedly, somatic ablation of Foxo1 in Neurog3(+) enteroendocrine progenitor cells gives rise to gut insulin-positive (Ins(+)) cells that express markers of mature β cells and secrete bioactive insulin as well as C-peptide in response to glucose and sulfonylureas. Lineage tracing experiments showed that gut Ins(+) cells arise cell autonomously from Foxo1-deficient cells. Inducible Foxo1 ablation in adult mice also resulted in the generation of gut Ins(+) cells. Following ablation by the β-cell toxin streptozotocin, gut Ins(+) cells regenerate and produce insulin, reversing hyperglycemia in mice. The data indicate that Neurog3(+) enteroendocrine progenitors require active Foxo1 to prevent differentiation into Ins(+) cells. Foxo1 ablation in gut epithelium may provide an approach to restore insulin production in type 1 diabetes.