Cancer mediates effector T cell dysfunction by targeting microRNAs and EZH2 via glycolysis restriction.

Aerobic glycolysis regulates T cell function. However, whether and how primary cancer alters T cell glycolytic metabolism and affects tumor immunity in cancer patients remains a question. Here we found that ovarian cancers imposed glucose restriction on T cells and dampened their function via maintaining high expression of microRNAs miR-101 and miR-26a, which constrained expression of the methyltransferase EZH2. EZH2 activated the Notch pathway by suppressing Notch repressors Numb and Fbxw7 via trimethylation of histone H3 at Lys27 and, consequently, stimulated T cell polyfunctional cytokine expression and promoted their survival via Bcl-2 signaling. Moreover, small hairpin RNA-mediated knockdown of human EZH2 in T cells elicited poor antitumor immunity. EZH2(+)CD8(+) T cells were associated with improved survival in patients. Together, these data unveil a metabolic target and mechanism of cancer immune evasion.

MS4A3 promotes differentiation in chronic myeloid leukemia by enhancing common ß-chain cytokine receptor endocytosis.

The chronic phase of chronic myeloid leukemia (CP-CML) is characterized by the excessive production of maturating myeloid cells. As CML stem/progenitor cells (LSPCs) are poised to cycle and differentiate, LSPCs must balance conservation and differentiation to avoid exhaustion, similar to normal hematopoiesis under stress. Since BCR-ABL1 tyrosine kinase inhibitors (TKIs) eliminate differentiating cells but spare BCR-ABL1-independent LSPCs, understanding the mechanisms that regulate LSPC differentiation may inform strategies to eliminate LSPCs. Upon performing a meta-analysis of published CML transcriptomes, we discovered that low expression of the MS4A3 transmembrane protein is a universal characteristic of LSPC quiescence, BCR-ABL1 independence, and transformation to blast phase (BP). Several mechanisms are involved in suppressing MS4A3, including aberrant methylation and a MECOM-C/EBPε axis. Contrary to previous reports, we find that MS4A3 does not function as a G1/S phase inhibitor but promotes endocytosis of common ß-chain (ßc) cytokine receptors upon GM-CSF/IL-3 stimulation, enhancing downstream signaling and cellular differentiation. This suggests that LSPCs downregulate MS4A3 to evade ßc cytokine-induced differentiation and maintain a more primitive, TKI-insensitive state. Accordingly, knockdown (KD) or deletion of MS4A3/Ms4a3 promotes TKI resistance and survival of CML cells ex vivo and enhances leukemogenesis in vivo, while targeted delivery of exogenous MS4A3 protein promotes differentiation. These data support a model in which MS4A3 governs response to differentiating myeloid cytokines, providing a unifying mechanism for the differentiation block characteristic of CML quiescence and BP-CML. Promoting MS4A3 reexpression or delivery of ectopic MS4A3 may help eliminate LSPCs in vivo.

Head and neck squamous cell carcinoma (HNSCC) in solid organ transplant recipients - Results from the scientific registry of transplant recipients (SRTR) database.

BACKGROUND: Solid organ transplant recipients have an elevated risk of cancer following organ transplantation than the age-adjusted general population. We assessed incidence of head and neck squamous cell carcinoma (HNSCC) in heart, lung, and liver recipients. BASIC PROCEDURES/METHODS: This retrospective cohort study included 124,966 patients from the United States Scientific Registry of Transplant Recipients (SRTR) database who received heart, lung, or liver transplantation between 1991 and 2010. Follow-up data were available until 2018. Patients with prevalent HNSCC at transplantation were excluded. Incident cases of HNSCC post organ transplantation were identified, and incidence rates (per 100,000 person-years) were reported by gender, race, organ type, year and age at organ transplantation. MAIN FINDINGS: The majority of patients received liver transplantation (58.64 %), followed by heart (28.64 %), and lung (12.72 %) transplantation. During follow-up, 4.14 % patients developed HNSCC. Overall incidence rate of HNSCC was 426.76 per 100,000 person-years. Male recipients had a higher HNSCC incidence rate than female recipients (571.8 and 177.0 per 100,000 person-years, respectively). Lung recipients had the highest overall HNSCC incidence rate (1273.6 per 100,000 person-years), followed by heart (644.2 per 100,000 person-years), and liver recipients (207.1 per 100,000 person-years). Overall, an increase in HNSCC incidence rate was observed with increase in age at organ transplantation. An increase in incidence rates of HNSCC over time was observed in lung recipients; however, incidence rates decreased over time in heart recipients. CONCLUSION: Solid organ transplant recipients have a high incidence of HNSCC following organ transplantation, and the incidence varies by type of organ received.

Identification of androgen response-related lncRNAs in prostate cancer.

BACKGROUND: Long noncoding RNAs (lncRNAs) are RNA molecules with over 200 nucleotides that do not code for proteins, but are known to be widely expressed and have key roles in gene regulation and cellular functions. They are also found to be involved in the onset and development of various cancers, including prostate cancer (PCa). Since PCa are commonly driven by androgen regulated signaling, mainly stimulated pathways, identification and determining the influence of lncRNAs in androgen response is useful and necessary. LncRNAs regulated by the androgen receptor (AR) can serve as potential biomarkers for PCa. In the present study, gene expression data analysis were performed to distinguish lncRNAs related to the androgen response pathway. METHODS AND RESULTS: We used publicly available RNA-sequencing and ChIP-seq data to identify lncRNAs that are associated with the androgen response pathway. Using Universal Correlation Coefficient (UCC) and Pearson Correlation Coefficient (PCC) analyses, we found 15 lncRNAs that have (a) highly correlated expression with androgen response genes in PCa and are (b) differentially expressed in the setting of treatment with an androgen agonist as well as antagonist compared to controls. Using publicly available ChIP-seq data, we investigated the role of androgen/AR axis in regulating expression of these lncRNAs. We observed AR binding in the promoter regions of 5 lncRNAs (MIR99AHG, DUBR, DRAIC, PVT1, and COLCA1), showing the direct influence of AR on their expression and highlighting their association with the androgen response pathway. CONCLUSION: By utilizing publicly available multiomics data and by employing in silico methods, we identified five candidate lncRNAs that are involved in the androgen response pathway. These lncRNAs should be investigated as potential biomarkers for PCa.

Nested and Large Nested Subtypes of Urothelial Carcinoma of the Upper Urinary Tract: A Multi-institutional Study.

Nested urothelial carcinoma (NUC) and large nested urothelial carcinoma (LNUC) of the upper urinary tract are exceedingly rare. This has contributed to the paucity of information regarding their clinicopathological and molecular characteristics. To address this knowledge gap, we explored the largest cohort to date of these rare tumors, comprising resection specimens of 10 LNUC and 7 NUC, from 7 participating institutions. Clinicopathological data were retrieved and documented. Whole exome sequencing and RNA sequencing were performed on the Illumina NovaSeq 6000 sequencer. The data generated were analyzed using the genome analysis toolkit pipeline. Somatic mutations were annotated using funcotator tool to identify pathogenic/likely pathogenic variants. Tumor mutational burden was calculated using python-based "pyTMB" tool. Microsatellite instability analysis was done using MSIsensor2 and the Idylla platform. Differential expression analysis of genes in LNUC and NUC along with mRNA expression-based molecular subtyping was performed by analyzing expression pattern of markers used in The Cancer Genome Atlas subclassification of bladder carcinoma. Both tumor types were more common in older males, were unifocal, and occurred more commonly mixed with minor components of predominantly conventional urothelial carcinoma. Overlying low-grade papillary urothelial carcinoma was significantly more common in LNUC (P = .034). On follow-up (LNUC: median, 10 months; range, 3-84 months; NUC: median, 9 months; range, 2-48 months), LNUC had better clinical outcomes (P = .031). Pathogenic mutations in FGFR3 and PIK3CA were significantly more common in LNUC (P = .049 and P = .044, respectively), with the latter present exclusively in LNUC. Seventy-five percent of the cases showed tumor mutational burden of <10, and all cases were microsatellite-stable. FGFR3 mutations were also more common in low-stage tumors. This study expands on the clinicopathological spectrum of NUC and LNUC of the upper urinary tract and is the first to comprehensively analyze the molecular profile of these tumors, highlighting pathogenic genetic alterations of potential therapeutic and prognostic value.

Developing 3D Organoid Raft Cultures from Patient-Derived Xenografts as Rapid Models to Screen Efficacy of Experimental Therapeutics.

Reliable preclinical models are needed for screening new cancer drugs. Thus, we developed an improved 3D tumor organoid model termed "organoid raft cultures" (ORCs). Development of ORCs involved culturing tumors ex vivo on collagen beds (boats) with grid supports to maintain their morphological structure. The ORCs were developed from patient-derived xenografts (PDXs) of colon cancers excised from immune-deficient mice (NOD/SCID/IL2Rgammanull). We utilized these new models to evaluate the efficacy of an investigational drug, Navitoclax (ABT-263). We tested the efficacy of ABT-263, an inhibitor of BCL-2 family proteins, in these ORCs derived from a PDX that showed high expression of antiapoptotic BCL2 family proteins (BCL-2, BCL-XL, and BCL-W). Hematoxylin and eosin staining evaluation of PDXs and corresponding ORCs indicated the retention of morphological and other histological integrity of ORCs. ORCs treated with ABT-263 showed decreased expression of antiapoptotic proteins (BCL2, BCL-XL and BCL-W) and increased proapoptotic proteins (BAX and PUMA), with concomitant activation of caspase 3. These studies support the usefulness of the ORCs, developed from PDXs, as an alternative to PDXs and as faster screening models.

14-3-3 proteins protect AMPK-phosphorylated ten-eleven translocation-2 (TET2) from PP2A-mediated dephosphorylation.

Ten-eleven translocation-2 (TET2) is a member of the methylcytosine dioxygenase family of enzymes and has been implicated in cancer and aging because of its role as a global epigenetic modifier. TET2 has a large N-terminal domain and a catalytic C-terminal region. Previous reports have demonstrated that the TET2 catalytic domain remains active independently of the N-terminal domain. As such, the function of the N terminus of this large protein remains poorly characterized. Here, using yeast two-hybrid screening, co-immunoprecipitation, and several biochemical assays, we found that several isoforms of the 14-3-3 family of proteins bind TET2. 14-3-3 proteins bound TET2 when it was phosphorylated at Ser-99. In particular, we observed that AMP-activated protein kinase-mediated phosphorylation at Ser-99 promotes TET2 stability and increases global DNA 5-hydroxymethylcytosine levels. The interaction of 14-3-3 proteins with TET2 protected the Ser-99 phosphorylation, and disruption of this interaction both reduced TET2 phosphorylation and decreased TET2 stability. Furthermore, we noted that protein phosphatase 2A can interact with TET2 and dephosphorylate Ser-99. Collectively, these results provide detailed insights into the role of the TET2 N-terminal domain in TET2 regulation. Moreover, they reveal the dynamic nature of TET2 protein regulation that could have therapeutic implications for disease states resulting from reduced TET2 levels or activity.

Characterization of the EZH2-MMSET histone methyltransferase regulatory axis in cancer.

Histone methyltransferases (HMTases), as chromatin modifiers, regulate the transcriptomic landscape in normal development as well in diseases such as cancer. Here, we molecularly order two HMTases, EZH2 and MMSET, that have established genetic links to oncogenesis. EZH2, which mediates histone H3K27 trimethylation and is associated with gene silencing, was shown to be coordinately expressed and function upstream of MMSET, which mediates H3K36 dimethylation and is associated with active transcription. We found that the EZH2-MMSET HMTase axis is coordinated by a microRNA network and that the oncogenic functions of EZH2 require MMSET activity. Together, these results suggest that the EZH2-MMSET HMTase axis coordinately functions as a master regulator of transcriptional repression, activation, and oncogenesis and may represent an attractive therapeutic target in cancer.

Fermentable fibers induce rapid macro- and micronutrient depletion in Toll-like receptor 5-deficient mice.

Functional fermentable fibers are considered essential for a healthy diet. Recently, we demonstrated that gut microbiota dysbiotic mice fed an inulin-containing diet (ICD) developed hepatocellular carcinoma (HCC) within 6 mo. In particular, a subset of Toll-like receptor 5-deficient (T5KO) mice prone to HCC exhibited rapid onset of hyperbilirubinemia (HB) and cholemia; these symptoms provide rationale that ICD induces cholestasis. Our objective in the present study was to determine whether inulin-fed T5KO-HB mice exhibit other known consequences of cholestasis, including essential fatty acid and fat-soluble vitamin deficiencies. Here, we measured hepatic fatty acids and serum vitamin A and D levels from wild-type (WT), T5KO low bilirubin (LB) and T5KO-HB mice fed ICD for 4 wk. Additionally, hepatic RNAseq and proteomics were performed to ascertain other metabolic alterations. Compared with WT and T5KO-LB, T5KO-HB mice exhibited steatorrhea, i.e., ~50% increase in fecal lipids. This could contribute to the significant reduction of linoleate in hepatic neutral lipids in T5KO-HB mice. Additionally, serum vitamins A and D were ~50% reduced in T5KO-HB mice, which was associated with metabolic compromises. Overall, our study highlights that fermentable fiber-induced cholestasis is further characterized by depletion of macro-and micronutrients.NEW & NOTEWORTHY Feeding a dietary, fermentable fiber diet to a subset of Toll-like receptor 5 deficient (T5KO) mice induces early onset hyperbilirubinemia and cholemia that later manifests to hepatocellular carcinoma (HCC). Our study highlights that fermentable fiber-induced cholestasis is characterized with modest macro- and micronutrient deficiencies that may further contribute to hepatic biliary disease. Compared with chemical induction, immunization, surgery, or genetic manipulation, these findings provide a novel approach to study the cholestatic subtype of HCC.

Genome-wide DNA methylation encodes cardiac transcriptional reprogramming in human ischemic heart failure.

Ischemic cardiomyopathy (ICM) is the clinical endpoint of coronary heart disease and a leading cause of heart failure. Despite growing demands to develop personalized approaches to treat ICM, progress is limited by inadequate knowledge of its pathogenesis. Since epigenetics has been implicated in the development of other chronic diseases, the current study was designed to determine whether transcriptional and/or epigenetic changes are sufficient to distinguish ICM from other etiologies of heart failure. Specifically, we hypothesize that genome-wide DNA methylation encodes transcriptional reprogramming in ICM. RNA-sequencing analysis was performed on human ischemic left ventricular tissue obtained from patients with end-stage heart failure, which enriched known targets of the polycomb methyltransferase EZH2 compared to non-ischemic hearts. Combined RNA sequencing and genome-wide DNA methylation analysis revealed a robust gene expression pattern consistent with suppression of oxidative metabolism, induced anaerobic glycolysis, and altered cellular remodeling. Lastly, KLF15 was identified as a putative upstream regulator of metabolic gene expression that was itself regulated by EZH2 in a SET domain-dependent manner. Our observations therefore define a novel role of DNA methylation in the metabolic reprogramming of ICM. Furthermore, we identify EZH2 as an epigenetic regulator of KLF15 along with DNA hypermethylation, and we propose a novel mechanism through which coronary heart disease reprograms the expression of both intermediate enzymes and upstream regulators of cardiac metabolism such as KLF15.

Characterization of glycine-N-acyltransferase like 1 (GLYATL1) in prostate cancer.

BACKGROUND: Recent microarray and sequencing studies of prostate cancer showed multiple molecular alterations during cancer progression. It is critical to evaluate these molecular changes to identify new biomarkers and targets. We performed analysis of glycine-N-acyltransferase like 1 (GLYATL1) expression in various stages of prostate cancer in this study and evaluated the regulation of GLYATL1 by androgen. METHOD: We performed in silico analysis of cancer gene expression profiling and transcriptome sequencing to evaluate GLYATL1 expression in prostate cancer. Furthermore, we performed immunohistochemistry using specific GLYATL1 antibody using high-density prostate cancer tissue microarray containing primary and metastatic prostate cancer. We also tested the regulation of GLYATL1 expression by androgen and ETS transcription factor ETV1. In addition, we performed RNA-sequencing of GLYATL1 modulated prostate cancer cells to evaluate the gene expression and changes in molecular pathways. RESULTS: Our in silico analysis of cancer gene expression profiling and transcriptome sequencing we revealed an overexpression of GLYATL1 in primary prostate cancer. Confirming these findings by immunohistochemistry, we show that GLYATL1 is overexpressed in primary prostate cancer compared with metastatic prostate cancer and benign prostatic tissue. Low-grade cancers had higher GLYATL1 expression compared to high-grade prostate tumors. Our studies showed that GLYATL1 is upregulated upon androgen treatment in LNCaP prostate cancer cells which harbors ETV1 gene rearrangement. Furthermore, ETV1 knockdown in LNCaP cells showed downregulation of GLYATL1 suggesting potential regulation of GLYATL1 by ETS transcription factor ETV1. Transcriptome sequencing using the GLYATL1 knockdown prostate cancer cell lines LNCaP showed regulation of multiple metabolic pathways. CONCLUSIONS: In summary, our study characterizes the expression of GLYATL1 in prostate cancer and explores the regulation of its regulation in prostate cancer showing role for androgen and ETS transcription factor ETV1. Future studies are needed to decipher the biological significance of these findings.

Large-scale profiling of serum metabolites in African American and European American patients with bladder cancer reveals metabolic pathways associated with patient survival.

BACKGROUND: African Americans (AAs) experience a disproportionally high rate of bladder cancer (BLCA) deaths even though their incidence rates are lower than those of other patient groups. Using a metabolomics approach, this study investigated how AA BLCA may differ molecularly from European Americans (EAs) BLCA, and it examined serum samples from patients with BLCA with the aim of identifying druggable metabolic pathways in AA patients. METHODS: Targeted metabolomics was applied to measure more than 300 metabolites in serum samples from 2 independent cohorts of EA and AA patients with BLCA and healthy EA and AA controls via liquid chromatography-mass spectrometry, and this was followed by the identification of altered metabolic pathways with a focus on AA BLCA. A subset of the differential metabolites was validated via absolute quantification with the Biocrates AbsoluteIDQ p180 kit. The clinical significance of the findings was further examined in The Cancer Genomic Atlas BLCA data set. RESULTS: Fifty-three metabolites, mainly related to amino acid, lipid, and nucleotide metabolism, were identified that showed significant differences in abundance between AA and EA BLCA. For example, the levels of taurine, glutamine, glutamate, aspartate, and serine were elevated in serum samples from AA patients versus EA patients. By mapping these metabolites to genes, this study identified significant relations with regulators of metabolism such as malic enzyme 3, prolyl 3-hydroxylase 2, and lysine demethylase 2A that predicted patient survival exclusively in AA patients with BLCA. CONCLUSIONS: This metabolic profile of serum samples might be used to assess risk progression in AA BLCA. These first-in-field findings describe metabolic alterations in AA BLCA and emphasize a potential biological basis for BLCA health disparities.

Wnt receptor Frizzled 8 is a target of ERG in prostate cancer.

Prostate cancer (PCa) is one of the most frequently diagnosed cancers among men. Many molecular changes have been detailed during PCa progression. The gene encoding the transcription factor ERG shows recurrent rearrangement, resulting in the overexpression of ERG in the majority of prostate cancers. Overexpression of ERG plays a critical role in prostate oncogenesis and development of metastatic disease. Among the downstream effectors of ERG, Frizzled family member FZD4 has been shown to be a target of ERG. Frizzled-8 (FZD8) has been shown to be involved in PCa bone metastasis. In the present study, we show that the expression of FZD8 is directly correlated with ERG expression in PCa. Furthermore, we show that ERG directly targets and activates FZD8 by binding to its promoter. This activation is specific to ETS transcription factor ERG and not ETV1. We propose that ERG overexpression in PCa leads to induction of Frizzled family member FZD8, which is known to activate the Wnt pathway. Taken together, these findings uncover a novel mechanism for PCa metastasis, and indicate that FZD8 may represent a potential therapeutic target for PCa.

Expression and Role of PAICS, a De Novo Purine Biosynthetic Gene in Prostate Cancer.

BACKGROUND: Our goal was to investigate de novo purine biosynthetic gene PAICS expression and evaluate its role in prostate cancer progression. METHODS: Next-generation sequencing, qRTPCR and immunoblot analysis revealed an elevated expression of a de novo purine biosynthetic gene, Phosphoribosylaminoimidazole Carboxylase, Phosphoribosylaminoimidazole Succinocarboxamide Synthetase (PAICS) in a progressive manner in prostate cancer. Functional analyses were performed using prostate cancer cell lines- DU145, PC3, LnCaP, and VCaP. The oncogenic properties of PAICS were studied both by transient and stable knockdown strategies, in vivo chicken chorioallantoic membrane (CAM) and murine xenograft models. Effect of BET bromodomain inhibitor JQ1 on the expression level of PAICS was also studied. RESULTS: Molecular staging of prostate cancer is important factor in effective diagnosis, prognosis and therapy. In this study, we identified a de novo purine biosynthetic gene; PAICS is overexpressed in PCa and its expression correlated with disease aggressiveness. Through several in vitro and in vivo functional studies, we show that PAICS is necessary for proliferation and invasion in prostate cancer cells. We identified JQ1, a BET bromodomain inhibitor previously implicated in regulating MYC expression and demonstrated role in prostate cancer, abrogates PAICS expression in several prostate cancer cells. Furthermore, we observe loss of MYC occupancy on PAICS promoter in presence of JQ1. CONCLUSIONS: Here, we report that evaluation of PAICS in prostate cancer progression and its role in prostate cancer cell proliferation and invasion and suggest it as a valid therapeutic target. We suggest JQ1, a BET-domain inhibitor, as possible therapeutic option in targeting PAICS in prostate cancer. Prostate 77:10-21, 2017. © 2016 Wiley Periodicals, Inc.

Genomic and Epigenomic Alterations in Cancer.

Multiple genetic and epigenetic events characterize tumor progression and define the identity of the tumors. Advances in high-throughput technologies, like gene expression profiling, next-generation sequencing, proteomics, and metabolomics, have enabled detailed molecular characterization of various tumors. The integration and analyses of these high-throughput data have unraveled many novel molecular aberrations and network alterations in tumors. These molecular alterations include multiple cancer-driving mutations, gene fusions, amplification, deletion, and post-translational modifications, among others. Many of these genomic events are being used in cancer diagnosis, whereas others are therapeutically targeted with small-molecule inhibitors. Multiple genes/enzymes that play a role in DNA and histone modifications are also altered in various cancers, changing the epigenomic landscape during cancer initiation and progression. Apart from protein-coding genes, studies are uncovering the critical regulatory roles played by noncoding RNAs and noncoding regions of the genome during cancer progression. Many of these genomic and epigenetic events function in tandem to drive tumor development and metastasis. Concurrent advances in genome-modulating technologies, like gene silencing and genome editing, are providing ability to understand in detail the process of cancer initiation, progression, and signaling as well as opening up avenues for therapeutic targeting. In this review, we discuss some of the recent advances in cancer genomic and epigenomic research.

Integrative genomics analysis reveals silencing of beta-adrenergic signaling by polycomb in prostate cancer.

The Polycomb group (PcG) protein EZH2 possesses oncogenic properties for which the underlying mechanism is unclear. We integrated in vitro cell line, in vivo tumor profiling, and genome-wide location data to nominate key targets of EZH2. One of the candidates identified was ADRB2 (Adrenergic Receptor, Beta-2), a critical mediator of beta-adrenergic signaling. EZH2 is recruited to the ADRB2 promoter and represses ADRB2 expression. ADRB2 inhibition confers cell invasion and transforms benign prostate epithelial cells, whereas ADRB2 overexpression counteracts EZH2-mediated oncogenesis. ADRB2 is underexpressed in metastatic prostate cancer, and clinically localized tumors that express lower levels of ADRB2 exhibit a poor prognosis. Taken together, we demonstrate the power of integrating multiple diverse genomic data to decipher targets of disease-related genes.

Metabolomic profiles delineate potential role for sarcosine in prostate cancer progression.

Multiple, complex molecular events characterize cancer development and progression. Deciphering the molecular networks that distinguish organ-confined disease from metastatic disease may lead to the identification of critical biomarkers for cancer invasion and disease aggressiveness. Although gene and protein expression have been extensively profiled in human tumours, little is known about the global metabolomic alterations that characterize neoplastic progression. Using a combination of high-throughput liquid-and-gas-chromatography-based mass spectrometry, we profiled more than 1,126 metabolites across 262 clinical samples related to prostate cancer (42 tissues and 110 each of urine and plasma). These unbiased metabolomic profiles were able to distinguish benign prostate, clinically localized prostate cancer and metastatic disease. Sarcosine, an N-methyl derivative of the amino acid glycine, was identified as a differential metabolite that was highly increased during prostate cancer progression to metastasis and can be detected non-invasively in urine. Sarcosine levels were also increased in invasive prostate cancer cell lines relative to benign prostate epithelial cells. Knockdown of glycine-N-methyl transferase, the enzyme that generates sarcosine from glycine, attenuated prostate cancer invasion. Addition of exogenous sarcosine or knockdown of the enzyme that leads to sarcosine degradation, sarcosine dehydrogenase, induced an invasive phenotype in benign prostate epithelial cells. Androgen receptor and the ERG gene fusion product coordinately regulate components of the sarcosine pathway. Here, by profiling the metabolomic alterations of prostate cancer progression, we reveal sarcosine as a potentially important metabolic intermediary of cancer cell invasion and aggressivity.

Inhibition of histone methylation arrests ongoing graft-versus-host disease in mice by selectively inducing apoptosis of alloreactive effector T cells.

Histone methylation is thought to be important for regulating Ag-driven T-cell responses. However, little is known about the effect of modulating histone methylation on inflammatory T-cell responses. We demonstrate that in vivo administration of the histone methylation inhibitor 3-deazaneplanocin A (DZNep) arrests ongoing GVHD in mice after allogeneic BM transplantation. DZNep caused selective apoptosis in alloantigen-activated T cells mediating host tissue injury. This effect was associated with the ability of DZNep to selectively reduce trimethylation of histone H3 lysine 27, deplete the histone methyltransferase Ezh2 specific to trimethylation of histone H3 lysine 27, and activate proapoptotic gene Bim repressed by Ezh2 in antigenic-activated T cells. In contrast, DZNep did not affect the survival of alloantigen-unresponsive T cells in vivo and naive T cells stimulated by IL-2 or IL-7 in vitro. Importantly, inhibition of histone methylation by DZNep treatment in vivo preserved the antileukemia activity of donor T cells and did not impair the recovery of hematopoiesis and lymphocytes, leading to significantly improved survival of recipients after allogeneic BM transplantation. Our findings indicate that modulation of histone methylation may have significant implications in the development of novel approaches to treat ongoing GVHD and other T cell-mediated inflammatory disorders in a broad context.

Targeting of oncogenic AAA-ATPase TRIP13 reduces progression of pancreatic ductal adenocarcinoma.

Thyroid hormone receptor-interacting protein 13 (TRIP13) is involved in cancer progression, but its role in pancreatic ductal adenocarcinoma (PDAC) is unknown. Thus, we assessed the expression, functional role, and mechanism of action of TRIP13 in PDAC. We further examined the efficacy of TRIP13 inhibitor, DCZ0415, alone or in combination with gemcitabine on malignant phenotypes, tumor progression, and immune response. We found that TRIP13 was overexpressed in human PDACs relative to corresponding normal pancreatic tissues. TRIP13 knockdown or treatment of PDAC cells with DCZ0415 reduced proliferation and colony formation, and induced G2/M cell cycle arrest and apoptosis. Additionally, TRIP13 knockdown or targeting with DCZ0415 reduced the migration and invasion of PDAC cells by increasing E-cadherin and decreasing N-cadherin and vimentin. Pharmacologic targeting or silencing of TRIP13 also resulted in reduce expression of FGFR4 and STAT3 phosphorylation, and downregulation of the Wnt/ß-catenin pathway. In immunocompromised mouse models of PDAC, knockdown of TRIP13 or treatment with DCZ0415 reduced tumor growth and metastasis. In an immunocompetent syngeneic PDAC model, DCZ0415 treatment enhanced the immune response by lowering expression of PD1/PDL1, increasing granzyme B/perforin expression, and facilitating infiltration of CD3/CD4 T-cells. Further, DCZ0415 potentiated the anti-metastatic and anti-tumorigenic activities of gemcitabine by reducing proliferation and angiogenesis and by inducing apoptosis and the immune response. These preclinical findings show that TRIP13 is involved in PDAC progression and targeting of TRIP13 augments the anticancer effect of gemcitabine.

Combination of dual JAK/HDAC inhibitor with regorafenib synergistically reduces tumor growth, metastasis, and regorafenib-induced toxicity in colorectal cancer.

BACKGROUND: Treatment with regorafenib, a multiple-kinase inhibitor, to manage metastatic colorectal cancers (mCRCs) shows a modest improvement in overall survival but is associated with severe toxicities. Thus, to reduce regorafenib-induced toxicity, we used regorafenib at low concentration along with a dual JAK/HDAC small-molecule inhibitor (JAK/HDACi) to leverage the advantages of both JAK and HDAC inhibition to enhance antitumor activity. The therapeutic efficacy and safety of the combination treatment was evaluated with CRC models. METHODS: The cytotoxicity of JAK/HDACi, regorafenib, and their combination were tested with normal colonic and CRC cells exhibiting various genetic backgrounds. Kinomic, ATAC-seq, RNA-seq, cell cycle, and apoptosis analyses were performed to evaluate the cellular functions/molecular alterations affected by the combination. Efficacy of the combination was assessed using patient-derived xenograft (PDX) and experimental metastasis models of CRC. To evaluate the interplay between tumor, its microenvironment, and modulation of immune response, MC38 syngeneic mice were utilized. RESULTS: The combination therapy decreased cell viability; phosphorylation of JAKs, STAT3, EGFR, and other key kinases; and inhibited deacetylation of histone H3K9, H4K8, and alpha tubulin proteins. It induced cell cycle arrest at G0-G1 phase and apoptosis of CRC cells. Whole transcriptomic analysis showed that combination treatment modulated molecules involved in apoptosis, extracellular matrix-receptor interaction, and focal adhesion pathways. It synergistically reduces PDX tumor growth and experimental metastasis, and, in a syngeneic mouse model, the treatment enhances the antitumor immune response as evidenced by higher infiltration of CD45 and cytotoxic cells. Pharmacokinetic studies showed that combination increased the bioavailability of regorafenib. CONCLUSIONS: The combination treatment was more effective than with regorafenib or JAK/HDACi alone, and had minimal toxicity. A clinical trial to evaluate this combination for treatment of mCRCs is warranted.