The potential of retinoids for combination therapy of lung cancer: Updates and future directions.

Lung cancer is the most common cancer-related death worldwide. Natural compounds have shown high biological and pharmaceutical relevance as anticancer agents. Retinoids are natural derivatives of vitamin A having many regulatory functions in the human body, including vision, cellular proliferation and differentiation, and activation of tumour suppressor genes. Retinoic acid (RA) is a highly active retinoid isoform with promising anti-lung cancer activity. The abnormal expression of retinoid receptors is associated with loss of anticancer activities and acquired resistance to RA in lung cancer. The preclinical promise has not translated to the general clinical utility of retinoids for lung cancer patients, especially those with a history of smoking. Newer retinoid nano-formulations and the combinatorial use of retinoids has been associated with lower toxicity and more favorably efficacy in both the preclinical and clinical settings. Here, we highlight epidemiological and clinical therapeutic studies involving retinoids and lung cancer. We also discuss the biological actions of retinoids in lung cancer, which include effects on cancer stem cell differentiation, angiogenesis, metastasis, and proliferative. We suggest that the use of retinoids in combination with conventional and targeted anticancer agents will broaden the utility of these potent anticancer compounds in the lung cancer clinic.

Perfluorooctanesulfonic Acid Alters Pro-Cancer Phenotypes and Metabolic and Transcriptional Signatures in Testicular Germ Cell Tumors.

The potential effects of poly- and perfluoroalkyl substances (PFAS) are a recently emergent human and environmental health concern. There is a consistent link between PFAS exposure and cancer, but the mechanisms are poorly understood. Although epidemiological evidence supporting PFAS exposure and cancer in general is conflicting, there is relatively strong evidence linking PFAS and testicular germ cell tumors (TGCTs). However, no mechanistic studies have been performed to date concerning PFAS and TGCTs. In this report, the effects of the legacy PFAS perfluorooctanesulfonic acid (PFOS) and the newer "clean energy" PFAS lithium bis(trifluoromethylsulfonyl)imide (LiTFSi, called HQ-115), on the tumorigenicity of TGCTs in mice, TGCT cell survival, and metabolite production, as well as gene regulation were investigated. In vitro, the proliferation and survival of both chemo-sensitive and -resistant TGCT cells were minimally affected by a wide range of PFOS and HQ-115 concentrations. However, both chemicals promoted the growth of TGCT cells in mouse xenografts at doses consistent with human exposure but induced minimal acute toxicity, as assessed by total body, kidney, and testis weight. PFOS, but not HQ-115, increased liver weight. Transcriptomic alterations of PFOS-exposed normal mouse testes were dominated by cancer-related pathways and gene expression alterations associated with the H3K27me3 polycomb pathway and DNA methylation, epigenetic pathways that were previously showed to be critical for the survival of TGCT cells after cisplatin-based chemotherapy. Similar patterns of PFOS-mediated gene expression occurred in PFOS-exposed cells in vitro. Metabolomic studies revealed that PFOS also altered metabolites associated with steroid biosynthesis and fatty acid metabolism in TGCT cells, consistent with the proposed ability of PFAS to mimic fatty acid-based ligands controlling lipid metabolism and the proposed role of PFAS as endocrine disrupters. Our data, is the first cell and animal based study on PFAS in TGCTs, support a pro-tumorigenic effect of PFAS on TGCT biology and suggests epigenetic, metabolic, and endocrine disruption as potential mechanisms of action that are consistent with the non-mutagenic nature of the PFAS class.

G0S2 promotes antiestrogenic and pro-migratory responses in ER+ and ER- breast cancer cells.

G0/G1 switch gene 2 (G0S2) is known to inhibit lipolysis by inhibiting adipose triglyceride lipase (ATGL). In this report, we dissect the role of G0S2 in ER+ versus ER- breast cancer. Overexpression of G0S2 in ER- cells increased cell proliferation, while G0S2 overexpression in ER+ cells decreased cell proliferation. Transcriptome analysis revealed that G0S2 mediated distinct but overlapping transcriptional responses in ER- and ER+ cells. G0S2 reduced genes associated with an epithelial phenotype, especially in ER- cells, including CDH1, ELF3, STEAP4 and TACSTD2, suggesting promotion of the epithelial-mesenchymal transition (EMT). G0S2 also repressed estrogen signaling and estrogen receptor target gene signatures, especially in ER+ cells, including TFF1 and TFF3. In addition, G0S2 overexpression increased cell migration in ER- cells and increased estrogen deprivation sensitivity in ER+ cells. Interestingly, two genes downstream of ATGL in fat utilization and very important in steroid hormone biosynthesis, HMGCS1 and HMGCS2, were downregulated in G0S2 overexpressing ER+ cells. In addition, HSD17B11, a gene that converts estradiol to its less estrogenic derivative, estrone, was highly upregulated in G0S2 overexpressing ER+ cells, suggesting G0S2 overexpression has a negative effect on estradiol production and maintenance. High expression of G0S2 and HSD17B11 was associated with improved relapse-free survival in breast cancer patients while high expression of HMGSC1 was associated with poor survival. Finally, we deleted G0S2 in breast cancer-prone MMTV-PyMT mice. Our data indicates a complex role for G0S2 in breast cancer, dependent on ER status, that may be partially mediated by suppression of the estrogen signaling pathway.

Serine/threonine kinase 17A is a novel p53 target gene and modulator of cisplatin toxicity and reactive oxygen species in testicular cancer cells.

Testicular cancer is highly curable with cisplatin-based therapy, and testicular cancer-derived human embryonal carcinoma (EC) cells undergo a p53-dominant transcriptional response to cisplatin. In this study, we have discovered that a poorly characterized member of the death-associated protein family of serine/threonine kinases, STK17A (also called DRAK1), is a novel p53 target gene. Cisplatin-mediated induction of STK17A in the EC cell line NT2/D1 was prevented with p53 siRNA. Furthermore, STK17A was induced with cisplatin in HCT116 and MCF10A cells but to a much lesser extent in isogenic p53-suppressed cells. A functional p53 response element that binds endogenous p53 in a cisplatin-dependent manner was identified 5 kb upstream of the first coding exon of STK17A. STK17A is not present in the mouse genome, but the closely related gene STK17B is induced with cisplatin in mouse NIH3T3 cells, although this induction is p53-independent. Interestingly, in human cells containing both STK17A and STK17B, only STK17A is induced with cisplatin. Knockdown of STK17A conferred resistance to cisplatin-induced growth suppression and apoptotic cell death in EC cells. This was associated with the up-regulation of detoxifying and antioxidant genes, including metallothioneins MT1H, MT1M, and MT1X that have previously been implicated in cisplatin resistance. In addition, knockdown of STK17A resulted in decreased cellular reactive oxygen species, whereas STK17A overexpression increased reactive oxygen species. In summary, we have identified STK17A as a novel direct target of p53 and a modulator of cisplatin toxicity and reactive oxygen species in testicular cancer cells.

The Relationship between Typical Environmental Endocrine Disruptors and Kidney Disease.

Endocrine disrupting chemicals (EDCs) are exogenous substances that alter the endocrine function of an organism, to result in adverse effects on growth and development, metabolism, and reproductive function. The kidney is one of the most important organs in the urinary system and an accumulation point. Studies have shown that EDCs can cause proteinuria, affect glomeruli and renal tubules, and even lead to diabetes and renal fibrosis in animal and human studies. In this review, we discuss renal accumulation of select EDCs such as dioxins, per- and polyfluoroalkyl substances (PFAS), bisphenol A (BPA), and phthalates, and delineate how exposures to such EDCs cause renal lesions and diseases, including cancer. The regulation of typical EDCs with specific target genes and the activation of related pathways are summarized.

Nephrotoxicity of perfluorooctane sulfonate (PFOS)-effect on transcription and epigenetic factors.

Perfluorooctane sulfonate (PFOS) is a widespread persistent environmental pollutant implicated in nephrotoxicity with altered metabolism, carcinogenesis, and fibrosis potential. We studied the underlying epigenetic mechanism involving transcription factors of PFOS-induced kidney injury. A 14-day orally dosed mouse model was chosen to study acute influences in vivo. Messenger RNA expression analysis and gene set enrichment analysis were performed to elucidate the relationship between epigenetic regulators, transcription factors, kidney disease, and metabolism homeostasis. PFOS was found to accumulate in mouse kidney in a dose-dependent manner. Kidney injury markers Acta2 and Bcl2l1 increased in expression significantly. Transcription factors, including Nef2l2, Hes1, Ppara, and Ppard, were upregulated, while Smarca2 and Pparg were downregulated. Furthermore, global DNA methylation levels decreased and the gene expression of histone demethylases Kdm1a and Kdm4c were upregulated. Our work implicates PFOS-induced gene expression alterations in epigenetics, transcription factors, and kidney biomarkers with potential implications for kidney fibrosis and kidney carcinogenesis. Future experiments can focus on epigenetic mechanisms to establish a panel of PFOS-induced biomarkers for nephrotoxicity evaluation.

Toward a Mechanistic Understanding of Poly- and Perfluoroalkylated Substances and Cancer.

Poly- and perfluoroalkylated substances (PFAS) are chemicals that persist and bioaccumulate in the environment and are found in nearly all human populations through several routes of exposure. Human occupational and community exposure to PFAS has been associated with several cancers, including cancers of the kidney, testis, prostate, and liver. While evidence suggests that PFAS are not directly mutagenic, many diverse mechanisms of carcinogenicity have been proposed. In this mini-review, we organize these mechanisms into three major proposed pathways of PFAS action-metabolism, endocrine disruption, and epigenetic perturbation-and discuss how these distinct but interdependent pathways may explain many of the proposed pro-carcinogenic effects of the PFAS class of environmental contaminants. Notably, each of the pathways is predicted to be highly sensitive to the dose and window of exposure which may, in part, explain the variable epidemiologic and experimental evidence linking PFAS and cancer. We highlight testicular and prostate cancer as models to validate this concept.

Reciprocal epigenetic remodeling controls testicular cancer hypersensitivity to hypomethylating agents and chemotherapy.

Testicular germ cell tumors (TGCTs) are aggressive but sensitive to cisplatin-based chemotherapy. Alternative therapies are needed for tumors refractory to cisplatin with hypomethylating agents providing one possibility. The mechanisms of cisplatin hypersensitivity and resistance in TGCTs remain poorly understood. Recently, it has been shown that TGCTs, even those resistant to cisplatin, are hypersensitive to very low doses of hypomethylating agents including 5-aza deoxy-cytosine (5-aza) and guadecitabine. We undertook a pharmacogenomic approach in order to better understand mechanisms of TGCT hypomethylating agent hypersensitivity by generating a panel of acquired 5-aza-resistant TGCT cells and contrasting these to previously generated acquired isogenic cisplatin-resistant cells from the same parent. Interestingly, there was a reciprocal relationship between cisplatin and 5-aza sensitivity, with cisplatin resistance associated with increased sensitivity to 5-aza and 5-aza resistance associated with increased sensitivity to cisplatin. Unbiased transcriptome analysis revealed 5-aza-resistant cells strongly downregulated polycomb target gene expression, the exact opposite of the finding for cisplatin-resistant cells, which upregulated polycomb target genes. This was associated with a dramatic increase in H3K27me3 and decrease in DNMT3B levels in 5-aza-resistant cells, the exact opposite changes seen in cisplatin-resistant cells. Evidence is presented that reciprocal regulation of polycomb and DNMT3B may be initiated by changes in DNMT3B levels as DNMT3B knockdown alone in parental cells resulted in increased expression of H3K27me3, EZH2, and BMI1, conferred 5-aza resistance and cisplatin sensitization, and mediated genome-wide repression of polycomb target gene expression. Finally, genome-wide analysis revealed that 5-aza-resistant, cisplatin-resistant, and DNMT3B-knockdown cells alter the expression of a common set of polycomb target genes. This study highlights that reciprocal epigenetic changes mediated by DNMT3B and polycomb may be a key driver of the unique cisplatin and 5-aza hypersensitivity of TGCTs and suggests that distinct epigenetic vulnerabilities may exist for pharmacological targeting of TGCTs.

The nuclear receptor TLX (NR2E1) inhibits growth and progression of triple- negative breast cancer.

Development of targeted therapies will be a critical step towards reducing the mortality associated with triple-negative breast cancer (TNBC). To achieve this, we searched for targets that met three criteria: (1) pharmacologically targetable, (2) expressed in TNBC, and (3) expression is prognostic in TNBC patients. Since nuclear receptors have a well-defined ligand-binding domain and are thus highly amenable to small-molecule intervention, we focused on this class of protein. Our analysis identified TLX (NR2E1) as a candidate. Specifically, elevated tumoral TLX expression was associated with prolonged recurrence-free survival and overall survival for breast cancer patients with either estrogen receptor alpha (ERα)-negative or basal-like tumors. Using two TNBC cell lines, we found that stable overexpression of TLX impairs in vitro proliferation. RNA-Seq analysis revealed that TLX reduced the expression of genes implicated in epithelial-mesenchymal transition (EMT), a cellular program known to drive metastatic progression. Indeed, TLX overexpression significantly decreased cell migration and invasion, and robustly decreased the metastatic capacity of TNBC cells in murine models. We identify SERPINB2 as a likely mediator of these effects. Taken together, our work indicates that TLX impedes the progression of TNBC. Several ligands have been shown to regulate the transcriptional activity of TLX, providing a framework for the future development of this receptor for therapeutic intervention.

Between a Rock and a Hard Place: An Epigenetic-Centric View of Testicular Germ Cell Tumors.

Compared to many common solid tumors, the main genetic drivers of most testicular germ cell tumors (TGCTs) are unknown. Decades of focus on genomic alterations in TGCTs including awareness of a near universal increase in copies of chromosome 12p have failed to uncover exceptional driver genes, especially in genes that can be targeted therapeutically. Thus far, TGCT patients have missed out on the benefits of targeted therapies available to treat most other malignancies. In the past decade there has been a greater appreciation that epigenetics may play an especially prominent role in TGCT etiology, progression, and hypersensitivity to conventional chemotherapy. While genetics undoubtedly plays a role in TGCT biology, this mini-review will focus on the epigenetic "states" or features of testicular cancer, with an emphasis on DNA methylation, histone modifications, and miRNAs associated with TGCT susceptibility, initiation, progression, and response to chemotherapy. In addition, we comment on the current status of epigenetic-based therapy and epigenetic biomarker development for TGCTs. Finally, we suggest a unifying "rock and a hard place" or "differentiate or die" model where the tumorigenicity and curability of TGCTs are both dependent on common but still ill-defined epigenetic states.

Per- and Polyfluoroalkyl Substance Exposure Combined with High-Fat Diet Supports Prostate Cancer Progression.

Per- and polyfluoroalkyl substances (PFAS) are synthetic chemicals utilized in various industrial settings and include products such as flame retardants, artificial film-forming foams, cosmetics, and non-stick cookware, among others. Epidemiological studies suggest a link between increased blood PFAS levels and prostate cancer incidence, but the mechanism through which PFAS impact cancer development is unclear. To investigate the link between PFAS and prostate cancer, we evaluated the impact of metabolic alterations resulting from a high-fat diet combined with PFAS exposure on prostate tumor progression. We evaluated in vivo prostate cancer xenograft models exposed to perfluorooctane sulfonate (PFOS), a type of PFAS compound, and different diets to study the effects of PFAS on prostate cancer progression and metabolic activity. Metabolomics and transcriptomics were used to understand the metabolic landscape shifts upon PFAS exposure. We evaluated metabolic changes in benign or tumor cells that lead to epigenomic reprogramming and altered signaling, which ultimately increase tumorigenic risk and tumor aggressiveness. Our studies are the first in the field to provide new and clinically relevant insights regarding novel metabolic and epigenetic states as well as to support the future development of effective preventative and therapeutic strategies for PFAS-induced prostate cancers. Our findings enhance understanding of how PFAS synergize with high-fat diets to contribute to prostate cancer development and establish an important basis to mitigate PFAS exposure.

Hypermethylation and global remodelling of DNA methylation is associated with acquired cisplatin resistance in testicular germ cell tumours.

Testicular germ cell tumours (TGCTs) respond well to cisplatin-based therapy. However, cisplatin resistance and poor outcomes do occur. It has been suggested that a shift towards DNA hypermethylation mediates cisplatin resistance in TGCT cells, although there is little direct evidence to support this claim. Here we utilized a series of isogenic cisplatin-resistant cell models and observed a strong association between cisplatin resistance in TGCT cells and a net increase in global CpG and non-CpG DNA methylation spanning regulatory, intergenic, genic and repeat elements. Hypermethylated loci were significantly enriched for repressive DNA segments, CTCF and RAD21 sites and lamina associated domains, suggesting that global nuclear reorganization of chromatin structure occurred in resistant cells. Hypomethylated CpG loci were significantly enriched for EZH2 and SUZ12 binding and H3K27me3 sites. Integrative transcriptome and methylome analyses showed a strong negative correlation between gene promoter and CpG island methylation and gene expression in resistant cells and a weaker positive correlation between gene body methylation and gene expression. A bidirectional shift between gene promoter and gene body DNA methylation occurred within multiple genes that was associated with upregulation of polycomb targets and downregulation of tumour suppressor genes. These data support the hypothesis that global remodelling of DNA methylation is a key factor in mediating cisplatin hypersensitivity and chemoresistance of TGCTs and furthers the rationale for hypomethylation therapy for refractory TGCT patients.

A phase 1 study of combined guadecitabine and cisplatin in platinum refractory germ cell cancer.

PURPOSE: Germ cell tumors (GCTs) are cured with therapy based on cisplatin, although a clinically significant number of patients are refractory and die of progressive disease. Based on preclinical studies indicating that refractory testicular GCTs are hypersensitive to hypomethylating agents (HMAs), we conducted a phase I trial combining the next-generation HMA guadecitabine (SGI-110) with cisplatin in recurrent, cisplatin-resistant GCT patients. METHODS: Patients with metastatic GCTs were treated for five consecutive days with guadecitabine followed by cisplatin on day 8, for a 28-day cycle for up to six cycles. The primary endpoint was safety and toxicity including dose-limiting toxicity (DLT) and maximum tolerated dose (MTD). RESULTS: The number of patients enrolled was 14. The majority of patients were heavily pretreated. MTD was determined to be 30 mg/m2 guadecitabine followed by 100 mg/m2 cisplatin. The major DLTs were neutropenia and thrombocytopenia. Three patients had partial responses by RECIST criteria, two of these patients, including one with primary mediastinal disease, completed the study and qualified as complete responses by serum tumor marker criteria with sustained remissions of 5 and 13 months and survival of 16 and 26 months, respectively. The overall response rate was 23%. Three patients also had stable disease indicating a clinical benefit rate of 46%. CONCLUSIONS: The combination of guadecitabine and cisplatin was tolerable and demonstrated activity in patients with platinum refractory germ cell cancer.

Wnt pathway reprogramming during human embryonal carcinoma differentiation and potential for therapeutic targeting.

BACKGROUND: Testicular germ cell tumors (TGCTs) are classified as seminonas or non-seminomas of which a major subset is embryonal carcinoma (EC) that can differentiate into diverse tissues. The pluripotent nature of human ECs resembles that of embryonic stem (ES) cells. Many Wnt signalling species are regulated during differentiation of TGCT-derived EC cells. This study comprehensively investigated expression profiles of Wnt signalling components regulated during induced differentiation of EC cells and explored the role of key components in maintaining pluripotency. METHODS: Human embryonal carcinoma cells were stably infected with a lentiviral construct carrying a canonical Wnt responsive reporter to assess Wnt signalling activity following induced differentiation. Cells were differentiated with all-trans retinoic acid (RA) or by targeted repression of pluripotency factor, POU5F1. A Wnt pathway real-time-PCR array was used to evaluate changes in gene expression as cells differentiated. Highlighted Wnt pathway genes were then specifically repressed using siRNA or stable shRNA and transfected EC cells were assessed for proliferation, differentiation status and levels of core pluripotency genes. RESULTS: Canonical Wnt signalling activity was low basally in undifferentiated EC cells, but substantially increased with induced differentiation. Wnt pathway gene expression levels were compared during induced differentiation and many components were altered including ligands (WNT2B), receptors (FZD5, FZD6, FZD10), secreted inhibitors (SFRP4, SFRP1), and other effectors of Wnt signalling (FRAT2, DAAM1, PITX2, Porcupine). Independent repression of FZD5, FZD7 and WNT5A using transient as well as stable methods of RNA interference (RNAi) inhibited cell growth of pluripotent NT2/D1 human EC cells, but did not appreciably induce differentiation or repress key pluripotency genes. Silencing of FZD7 gave the greatest growth suppression in all human EC cell lines tested including NT2/D1, NT2/D1-R1, Tera-1 and 833K cells. CONCLUSION: During induced differentiation of human EC cells, the Wnt signalling pathway is reprogrammed and canonical Wnt signalling induced. Specific species regulating non-canonical Wnt signalling conferred growth inhibition when targeted for repression in these EC cells. Notably, FZD7 repression significantly inhibited growth of human EC cells and is a promising therapeutic target for TGCTs.

Mechanisms of cisplatin sensitivity and resistance in testicular germ cell tumors.

Testicular germ cell tumors (TGCTs) are a cancer pharmacology success story with a majority of patients cured even in the highly advanced and metastatic setting. Successful treatment of TGCTs is primarily due to the exquisite responsiveness of this solid tumor to cisplatin-based therapy. However, a significant percentage of patients are, or become, refractory to cisplatin and die from progressive disease. Mechanisms for both clinical hypersensitivity and resistance have largely remained a mystery despite the promise of applying lessons to the majority of solid tumors that are not curable in the metastatic setting. Recently, this promise has been heightened by the realization that distinct (and perhaps pharmacologically replicable) epigenetic states, rather than fixed genetic alterations, may play dominant roles in not only TGCT etiology and progression but also their curability with conventional chemotherapies. In this review, it discusses potential mechanisms of TGCT cisplatin sensitivity and resistance to conventional chemotherapeutics.

Epigenetic Remodeling through Downregulation of Polycomb Repressive Complex 2 Mediates Chemotherapy Resistance in Testicular Germ Cell Tumors.

A greater understanding of the hypersensitivity and curability of testicular germ cell tumors (TGCTs) has the potential to inform strategies to sensitize other solid tumors to conventional chemotherapies. The mechanisms of cisplatin hypersensitivity and resistance in embryonal carcinoma (EC), the stem cells of TGCTs, remain largely undefined. To study the mechanisms of cisplatin resistance we generated a large panel of independently derived, acquired resistant clones from three distinct parental EC models employing a protocol designed to match standard of care regimens of TGCT patients. Transcriptomics revealed highly significant expression changes shared between resistant cells regardless of their parental origin. This was dominated by a highly significant enrichment of genes normally repressed by H3K27 methylation and the polycomb repressive complex 2 (PRC2) which correlated with a substantial decrease in global H3K27me3, H2AK119 ubiquitination, and expression of BMI1. Importantly, repression of H3K27 methylation with the EZH2 inhibitor GSK-126 conferred cisplatin resistance to parental cells while induction of H3K27 methylation with the histone lysine demethylase inhibitor GSK-J4 resulted in increased cisplatin sensitivity to resistant cells. A gene signature based on H3K27me gene enrichment was associated with an increased rate of recurrent/progressive disease in testicular cancer patients. Our data indicates that repression of H3K27 methylation is a mechanism of cisplatin acquired resistance in TGCTs and that restoration of PRC2 complex function is a viable approach to overcome treatment failure.

The direct p53 target gene, FLJ11259/DRAM, is a member of a novel family of transmembrane proteins.

The tumor suppressor p53 regulates diverse biological processes primarily via activation of downstream target genes. Even though many p53 target genes have been described, the precise mechanisms of p53 biological actions are uncertain. In previous work we identified by microarray analysis a candidate p53 target gene, FLJ11259/DRAM. In this report we have identified three uncharacterized human proteins with sequence homology to FLJ11259, suggesting that FLJ11259 is a member of a novel family of proteins with six transmembrane domains. Several lines of investigation confirm FLJ11259 is a direct p53 target gene. p53 siRNA prevented cisplatin-mediated up-regulation of FLJ11259 in NT2/D1 cells. Likewise in HCT116 p53+/+ cells and MCF10A cells, FLJ11259 is induced by cisplatin treatment but to a much lesser extent in isogenic p53-suppressed cells. A functional p53 response element was identified 22.3 kb upstream of the first coding exon of FLJ11259 and is shown to be active in reporter assays. In addition, chromatin immunoprecipitation assays indicate that p53 binds directly to this element in vivo and that binding is enhanced following cisplatin treatment. Confocal microscopy showed that an FLJ-GFP fusion protein localizes mainly in a punctate pattern in the cytoplasm. Overexpression studies in Cos-7, Saos2, and NT2/D1 cells suggest that FLJ11259 is associated with increased clonal survival. In summary, we have identified FLJ11259/DRAM as a p53-inducible member of a novel family of transmembrane proteins. FLJ11259/DRAM may be an important modulator of p53 responses in diverse tumor types.

Incorporating DNA Methyltransferase Inhibitors (DNMTis) in the Treatment of Genitourinary Malignancies: A Systematic Review.

Inhibition of DNA methyltransferases (DNMTs) has emerged as a novel treatment strategy in solid tumors. Aberrant hypermethylation in promoters of critical tumor suppressor genes is the basis for the idea that treatment with hypomethylating agents may lead to the restoration of a "normal" epigenome and produce clinically meaningful therapeutic outcomes. The aim of this review article is to summarize the current state of knowledge of DNMT inhibitors in the treatment of genitourinary malignancies. The efficacy of these agents in genitourinary malignancies was reported in a number of studies and suggests a role of induced DNA hypomethylation in overcoming resistance to conventional cytotoxic treatments. The clinical significance of these findings should be further investigated.

Selective repression of retinoic acid target genes by RIP140 during induced tumor cell differentiation of pluripotent human embryonal carcinoma cells.

BACKGROUND: The use of retinoids as anti-cancer agents has been limited due to resistance and low efficacy. The dynamics of nuclear receptor coregulation are incompletely understood. Cell-and context-specific activities of nuclear receptors may be in part due to distinct coregulator complexes recruited to distinct subsets of target genes. RIP140 (also called NRIP1) is a ligand-dependent corepressor that is inducible with retinoic acid (RA). We had previously shown that RIP140 limits RA induced tumor cell differentiation of embryonal carcinoma; the pluriopotent stem cells of testicular germ cell tumors. This implies that RIP140 represses key genes required for RA-mediated tumor cell differentiation. Identification of these genes would be of considerable interest. RESULTS: To begin to address this issue, microarray technology was employed to elucidate in a de novo fashion the global role of RIP140 in RA target gene regulation of embryonal carcinoma. Subclasses of genes were affected by RIP140 in distinct manners.Interestingly, approximately half of the RA-dependent genes were unaffected by RIP140. Hence, RIP140 appears to discriminate between different classes of RA target genes. In general, RIP140-dependent gene expression was consistent with RIP140 functioning to limit RA signaling and tumor cell differentiation. Few if any genes were regulated in a manner to support a role for RIP140 in "active repression". We also demonstrated that RIP140 silencing sensitizes embryonal carcinoma cells to low doses of RA. CONCLUSION: Together the data demonstrates that RIP140 has profound effects on RA-mediated gene expression in this cancer stem cell model. The RIP140-dependent RA target genes identified here may be particularly important in mediating RA-induced tumor cell differentiation and the findings suggest that RIP140 may be an attractive target to sensitize tumor cells to retinoid-based differentiation therapy. We discuss these data in the context of proposed models of RIP140-mediated repression.

Repression of cyclin D1 as a target for germ cell tumors.

Metastatic germ cell tumors (GCT) are curable, however GCTs refractory to cisplatin-based chemotherapy have a poor prognosis. This study explores D-type cyclins as molecular targets in GCTs because all-trans-retinoic acid (RA)-mediated differentiation of the human embryonal carcinoma (EC) cell line NT2/D1 is associated with G1 cell cycle arrest and proteasomal degradation of cyclin D1. RA effects on D-type cyclins are compared in human EC cells that are RA sensitive or dually RA and cisplatin resistant (NT2/D1-R1) and in clinical GCTs that have both EC and mature teratoma components. Notably, GCT differentiation was associated with reduced cyclin D1 but increased cyclin D3 expression. RA was shown here to repress cyclin D1 through a transcriptional mechanism in addition to causing its degradation. The siRNA-mediated repression of individual cyclin D species resulted in growth inhibition in both RA sensitive and resistant EC cells. Only repression of cyclin D1 occurred in vitro and when clinical GCTs mature, implicating cyclin D1 as a molecular therapeutic target. To confirm this, the EGFR-tyrosine kinase inhibitor, Erlotinib, was used to repress cyclin D1. This inhibited proliferation in RA and cisplatin sensitive and resistant EC cells. Taken together, these findings implicate cyclin D1 targeting agents for the treatment of GCTs.