Collateral damage of NUDT15 deficiency in cancer provides a cancer pharmacogenetic therapeutic window with thiopurines.

Genome instability is a hallmark of cancer and are driven by mutations in oncogenes and tumor suppressor genes. Despite successes seen with select targeted therapeutics, this type of personalized medicine is only beneficial for a small subpopulation of cancer patients who have one of a few actionable genetic changes. Most tumors also contain hundreds of passenger mutations that offered no fitness advantage or disadvantage during tumor evolution. Mutations in known pharmacogenetic (PGx) loci for which germline variants encode variability in drug response can cause somatically acquired drug sensitivity. The NUDT15 gene is a known PGx locus that participates in the rate-limiting metabolism of thiopurines. People with two defective germline alleles of NUDT15 are hypersensitive to the toxic effects of thiopurines. NUDT15 is located adjacent to the Retinoblastoma ( RB1 ) tumor suppressor gene, which often undergoes homozygous deletion in retinoblastomas and other epithelial cancers. We observed that RB1 undergoes homozygous deletions in 9.4% of prostate adenocarcinomas and 2.5% of ovarian cancers, and in nearly all of these cases NUDT15 is also lost. Moreover, 44% of prostate adenocarcinomas and over 60% of ovarian cancers have lost one allele of NUDT15, which predicts that a majority of all prostate and ovarian cancers have somatically acquired hypersensitivity to thiopurine treatment. We performed a retrospective analysis of >16,000 patients in the US Veterans Administration health care system and found concurrent xanthine oxidase inhibition (XOi) and thiopurine usage for non-cancer indications is significantly associated with reduced incidence of prostate cancer. The hazard ratio for the development of prostate cancer in patients treated with thiopurines and XOi was 0.562 (0.301-1.051) for the unmatched cohort and 0.389 (0.185-0.819) for the propensity score matched cohort. We experimentally depleted NUDT15 from ovarian and prostate cancer cell lines and observed a dramatic sensitization to thiopurine-induced and DNA damage-dependent toxicity. These results indicate that somatic loss of NUDT15 predicts therapeutic sensitivity to a low cost and well tolerated drug with a broad therapeutic window.

The synthetic food dye, Red 40, causes DNA damage, causes colonic inflammation, and impacts the microbiome in mice.

The incidence of colorectal cancer (CRC) among young people has been on the rise for the past four decades and its underlying causes are only just starting to be uncovered. Recent studies suggest that consuming ultra-processed foods and pro-inflammatory diets may be contributing factors. The increase in the use of synthetic food colors in such foods over the past 40 years, including the common synthetic food dye Allura Red AC (Red 40), coincides with the rise of early-onset colorectal cancer (EOCRC). As these ultra-processed foods are particularly appealing to children, there is a growing concern about the impact of synthetic food dyes on the development of CRC. Our study aimed to investigate the effects of Red 40 on DNA damage, the microbiome, and colonic inflammation. Despite a lack of prior research, high levels of human exposure to pro-inflammatory foods containing Red 40 highlight the urgency of exploring this issue. Our results show that Red 40 damages DNA both in vitro and in vivo and that consumption of Red 40 in the presence of a high-fat diet for 10 months leads to dysbiosis and low-grade colonic inflammation in mice. This evidence supports the hypothesis that Red 40 is a dangerous compound that dysregulates key players involved in the development of EOCRC.

Secretory pathway Ca2+-ATPase SPCA2 regulates mitochondrial respiration and DNA damage response through store-independent calcium entry.

A complex interplay between the extracellular space, cytoplasm and individual organelles modulates Ca2+ signaling to impact all aspects of cell fate and function. In recent years, the molecular machinery linking endoplasmic reticulum stores to plasma membrane Ca2+ entry has been defined. However, the mechanism and pathophysiological relevance of store-independent modes of Ca2+ entry remain poorly understood. Here, we describe how the secretory pathway Ca2+-ATPase SPCA2 promotes cell cycle progression and survival by activating store-independent Ca2+ entry through plasma membrane Orai1 channels in mammary epithelial cells. Silencing SPCA2 expression or briefly removing extracellular Ca2+ increased mitochondrial ROS production, DNA damage and activation of the ATM/ATR-p53 axis leading to G0/G1 phase cell cycle arrest and apoptosis. Consistent with these findings, SPCA2 knockdown confers redox stress and chemosensitivity to DNA damaging agents. Unexpectedly, SPCA2-mediated Ca2+ entry into mitochondria is required for optimal cellular respiration and the generation of mitochondrial membrane potential. In hormone receptor positive (ER+/PR+) breast cancer subtypes, SPCA2 levels are high and correlate with poor survival prognosis. We suggest that elevated SPCA2 expression could drive pro-survival and chemotherapy resistance in cancer cells, and drugs that target store-independent Ca2+ entry pathways may have therapeutic potential in treating cancer.

Serine-threonine Kinase Receptor-Associated Protein is a Critical Mediator of APC Mutation-Induced Intestinal Tumorigenesis Through a Feed-Forward Mechanism.

BACKGROUND & AIMS: Inactivation of the Apc gene is a critical early event in the development of sporadic colorectal cancer (CRC). Expression of serine-threonine kinase receptor-associated protein (STRAP) is elevated in CRCs and is associated with poor outcomes. We investigated the role of STRAP in Apc mutation-induced intestinal tumor initiation and progression. METHODS: We generated Strap intestinal epithelial knockout mice (StrapΔIEC) by crossing mice containing floxed alleles of Strap (Strapfl/fl) with Villin-Cre mice. Then we generated ApcMin/+;Strapfl/fl;Vill-Cre (ApcMin/+;StrapΔIEC) mice for RNA-sequencing analyses to determine the mechanism of function of STRAP. We used human colon cancer cell lines (DLD1, SW480, and HT29) and human and mouse colon tumor-derived organoids for STRAP knockdown and knockout and overexpression experiments. RESULTS: Strap deficiency extended the average survival of ApcMin/+ mice by 80 days and decreased the formation of intestinal adenomas. Expression profiling revealed that the intestinal stem cell signature, the Wnt/ß-catenin signaling, and the MEK/ERK pathway are down-regulated in Strap-deficient adenomas and intestinal organoids. Correlation studies suggest that these STRAP-associated oncogenic signatures are conserved across murine and human colon cancer. STRAP associates with MEK1/2, promotes binding between MEK1/2 and ERK1/2, and subsequently induces the phosphorylation of ERK1/2. STRAP activated Wnt/ß-catenin signaling through MEK/ERK-induced phosphorylation of LRP6. STRAP was identified as a target of mutated Apc and Wnt/ß-catenin signaling as chromatin immunoprecipitation and luciferase assays revealed putative binding sites of the ß-catenin/TCF4 complex on the Strap promoter. CONCLUSIONS: STRAP is a target of, and is required in, Apc mutation/deletion-induced intestinal tumorigenesis through a novel feed-forward STRAP/MEK-ERK/Wnt-ß-catenin/STRAP regulatory axis.

HPV-inactive cell populations arise from HPV16-transformed human keratinocytes after p53 knockout.

HPV-inactive head and neck and cervical cancers contain HPV DNA but do not express HPV E6/E7. HPV-positive primary head and neck tumors usually express E6/E7, however they may produce HPV-inactive metastases. These observations led to our hypothesis that HPV-inactive cancers begin as HPV-active lesions, losing dependence on E6/E7 expression during progression. Because HPV-inactive cervical cancers often have mutated p53, we investigated whether p53 loss may play a role in the genesis of HPV-inactive cancers. p53 knockout (p53-KO) by CRISPR-Cas9 resulted in a 5-fold reduction of E7 mRNA in differentiation-resistant HPV16 immortalized human keratinocytes (HKc/DR). E7 expression was restored by 5-Aza-2 deoxycytidine in p53 KO lines, suggesting a role of DNA methylation in this process. In-situ hybridization showed that p53 KO lines consist of mixed populations of E6/E7-positive and negative cells. Hence, loss of p53 predisposes HPV16 transformed cells to losing dependence on the continuous expression of HPV oncogenes for proliferation.

Publisher Correction: Early-onset colorectal cancer: initial clues and current views.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Targeting lysyl oxidase (LOX) overcomes chemotherapy resistance in triple negative breast cancer.

Chemoresistance is a major obstacle in triple negative breast cancer (TNBC), the most aggressive breast cancer subtype. Here we identify hypoxia-induced ECM re-modeler, lysyl oxidase (LOX) as a key inducer of chemoresistance by developing chemoresistant TNBC tumors in vivo and characterizing their transcriptomes by RNA-sequencing. Inhibiting LOX reduces collagen cross-linking and fibronectin assembly, increases drug penetration, and downregulates ITGA5/FN1 expression, resulting in inhibition of FAK/Src signaling, induction of apoptosis and re-sensitization to chemotherapy. Similarly, inhibiting FAK/Src results in chemosensitization. These effects are observed in 3D-cultured cell lines, tumor organoids, chemoresistant xenografts, syngeneic tumors and PDX models. Re-expressing the hypoxia-repressed miR-142-3p, which targets HIF1A, LOX and ITGA5, causes further suppression of the HIF-1α/LOX/ITGA5/FN1 axis. Notably, higher LOX, ITGA5, or FN1, or lower miR-142-3p levels are associated with shorter survival in chemotherapy-treated TNBC patients. These results provide strong pre-clinical rationale for developing and testing LOX inhibitors to overcome chemoresistance in TNBC patients.

PRDM16 suppresses HIF-targeted gene expression in kidney cancer.

Analysis of transcriptomic data demonstrates extensive epigenetic gene silencing of the transcription factor PRDM16 in renal cancer. We show that restoration of PRDM16 in RCC cells suppresses in vivo tumor growth. RNaseq analysis reveals that PRDM16 imparts a predominantly repressive effect on the RCC transcriptome including suppression of the gene encoding semaphorin 5B (SEMA5B). SEMA5B is a HIF target gene highly expressed in RCC that promotes in vivo tumor growth. Functional studies demonstrate that PRDM16's repressive properties, mediated by physical interaction with the transcriptional corepressors C-terminal binding proteins (CtBP1/2), are required for suppression of both SEMA5B expression and in vivo tumor growth. Finally, we show that reconstitution of RCC cells with a PRDM16 mutant unable to bind CtBPs nullifies PRDM16's effects on both SEMA5B repression and tumor growth suppression. Collectively, our data uncover a novel epigenetic basis by which HIF target gene expression is amplified in kidney cancer and a new mechanism by which PRDM16 exerts its tumor suppressive effects.

Early-onset colorectal cancer: initial clues and current views.

Over the past several decades, the incidence of early-onset colorectal cancer (EOCRC; in patients <50 years old) has increased at an alarming rate. Although robust and scientifically rigorous epidemiological studies have sifted out environmental elements linked to EOCRC, our knowledge of the causes and mechanisms of this disease is far from complete. Here, we highlight potential risk factors and putative mechanisms that drive EOCRC and suggest likely areas for fruitful research. In addition, we identify inconsistencies in the evidence implicating a strong effect of increased adiposity and suggest that certain behaviours (such as diet and stress) might place nonobese and otherwise healthy people at risk of this disease. Key risk factors are reviewed, including the global westernization of diets (usually involving a high intake of red and processed meats, high-fructose corn syrup and unhealthy cooking methods), stress, antibiotics, synthetic food dyes, monosodium glutamate, titanium dioxide, and physical inactivity and/or sedentary behaviour. The gut microbiota is probably at the crossroads of these risk factors and EOCRC. The time course of the disease and the fact that relevant exposures probably occur in childhood raise important methodological issues that are also discussed.

Spindle Assembly Checkpoint Inhibition Can Resensitize p53-Null Stem Cells to Cancer Chemotherapy.

TP53 mutations are common in most human cancers, but few therapeutic options for TP53-mutant tumors exist. To identify potential therapeutic options for cancer patients with TP53 mutations, we profiled 127 FDA-approved chemotherapy drugs against human embryonic stem cells (hESC) in which we engineered TP53 deletion by genome editing. We identified 27 cancer therapeutic drugs for which TP53 mutations conferred resistance; most of these drugs target DNA synthesis or topoisomerase and cause DNA damage. We then performed a genome-wide CRISPR/Cas9 knockout screen in the TP53-null hESC in the presence and absence of sublethal concentrations of cisplatin and identified 137 genes whose loss selectively resensitized the p53-null cells to this chemotherapeutic agent. Gene ontology classification of the resensitizing loci revealed significant overrepresentation of spindle checkpoint pathway genes. Moreover, we confirmed that targeting ZNF207/BuGZ sensitizes p53-null hESC to cisplatin. These data indicate that targeted inhibition of spindle assembly checkpoints (SAC) and chromosomal organizing centers may provide a way to treat p53-deficient cancer cells with standard chemotherapy drugs. Development of small-molecule inhibitors of SAC proteins may be a useful strategy for rescuing DNA-damaging chemotherapeutics in TP53-mutant cancers. SIGNIFICANCE: These findings show that inhibition of spindle assembly checkpoints and chromosomal organizing centers may provide a new way to treat p53-deficient cancer cells with standard chemotherapy drugs.

Myeloid-Derived Suppressor Cells Produce IL-10 to Elicit DNMT3b-Dependent IRF8 Silencing to Promote Colitis-Associated Colon Tumorigenesis.

IL-10 functions as a suppressor of colitis and colitis-associated colon cancer, but it is also a risk locus associated with ulcerative colitis. The mechanism underlying the contrasting roles of IL-10 in inflammation and colon cancer is unknown. We report here that inflammation induces the accumulation of CD11b+Gr1+ myeloid-derived suppressor cells (MDSCs) that express high levels of IL-10 in colon tissue. IL-10 induces the activation of STAT3 that directly binds to the Dnmt1 and Dnmt3b promoters to activate their expression, resulting in DNA hypermethylation at the Irf8 promoter to silence IRF8 expression in colon epithelial cells. Mice with Irf8 deleted in colonic epithelial cells exhibit significantly higher inflammation-induced tumor incidence. Human colorectal carcinomas have significantly higher DNMT1 and DNMT3b and lower IRF8 expression, and they exhibit significantly higher IRF8 promoter DNA methylation than normal colon. Our data identify the MDSC-IL-10-STAT3-DNMT3b-IRF8 pathway as a link between chronic inflammation and colon cancer initiation.

Biochemical and Epigenetic Insights into L-2-Hydroxyglutarate, a Potential Therapeutic Target in Renal Cancer.

PURPOSE: Elevation of L-2-hydroxylgutarate (L-2-HG) in renal cell carcinoma (RCC) is due in part to reduced expression of L-2-HG dehydrogenase (L2HGDH). However, the contribution of L-2-HG to renal carcinogenesis and insight into the biochemistry and targets of this small molecule remains to be elucidated. EXPERIMENTAL DESIGN: Genetic and pharmacologic approaches to modulate L-2-HG levels were assessed for effects on in vitro and in vivo phenotypes. Metabolomics was used to dissect the biochemical mechanisms that promote L-2-HG accumulation in RCC cells. Transcriptomic analysis was utilized to identify relevant targets of L-2-HG. Finally, bioinformatic and metabolomic analyses were used to assess the L-2-HG/L2HGDH axis as a function of patient outcome and cancer progression. RESULTS: L2HGDH suppresses both in vitro cell migration and in vivo tumor growth and these effects are mediated by L2HGDH's catalytic activity. Biochemical studies indicate that glutamine is the predominant carbon source for L-2-HG via the activity of malate dehydrogenase 2 (MDH2). Inhibition of the glutamine-MDH2 axis suppresses in vitro phenotypes in an L-2-HG-dependent manner. Moreover, in vivo growth of RCC cells with basal elevation of L-2-HG is suppressed by glutaminase inhibition. Transcriptomic and functional analyses demonstrate that the histone demethylase KDM6A is a target of L-2-HG in RCC. Finally, increased L-2-HG levels, L2HGDH copy loss, and lower L2HGDH expression are associated with tumor progression and/or worsened prognosis in patients with RCC. CONCLUSIONS: Collectively, our studies provide biochemical and mechanistic insight into the biology of this small molecule and provide new opportunities for treating L-2-HG-driven kidney cancers.

PRDM1 silences stem cell-related genes and inhibits proliferation of human colon tumor organoids.

PRDM1 is a tumor suppressor that plays an important role in B and T cell lymphomas. Our previous studies demonstrated that PRDM1ß is a p53-response gene in human colorectal cancer cells. However, the function of PRDM1ß in colorectal cancer cells and colon tumor organoids is not clear. Here we show that PRDM1ß is a p53-response gene in human colon organoids and that low PRDM1 expression predicts poor survival in colon cancer patients. We engineered PRDM1 knockouts and overexpression clones in RKO cells and characterized the PRDM1-dependent transcript landscapes, revealing that both the α and ß transcript isoforms repress MYC-response genes and stem cell-related genes. Finally, we show that forced expression of PRDM1 in human colon cancer organoids prevents the formation and growth of colon tumor organoids in vitro. These results suggest that p53 may exert tumor-suppressive effects in part through a PRDM1-dependent silencing of stem cell genes, depleting the size of the normal intestinal stem cell compartment in response to DNA damage.

Temporal Efficacy of a Sulforaphane-Based Broccoli Sprout Diet in Prevention of Breast Cancer through Modulation of Epigenetic Mechanisms.

Breast cancer is the most common cancer and the second leading cause of cancer-related death among women. An important risk factor for breast cancer is individual genetic background, which is initially generated early in human life, for example, during the processes of embryogenesis and fetal development in utero Bioactive dietary components such as sulforaphane (SFN), an isothiocyanate from cruciferous vegetables including broccoli sprouts (BSp), cabbage, and kale, has been shown to reduce the risk of developing many common cancers through regulation of epigenetic mechanisms. Our study indicates a prenatal/maternal BSp dietary treatment exhibited maximal preventive effects in inhibiting breast cancer development compared with postnatal early-life and adult BSp treatments in two transgenic mouse models that can develop breast cancer. Postnatal early-life BSp treatment starting prior to puberty onset showed protective effects in prevention of breast cancer but was not as effective as the prenatal/maternal BSp treatment. However, adulthood-administered BSp diet did not reduce mammary tumorigenesis. Our results suggest that the prenatal/maternal BSp bioactive natural plant product may impact early embryonic development by regulating global differential gene expression through affecting epigenetic profiles resulting in differential susceptibility to breast cancer later in life. These results suggest that a temporal exposure to epigenetic-modulating dietary components such as cruciferous vegetables could be a key factor for maximizing chemopreventive effects on human breast cancer. This study may lead to translational breast cancer chemopreventive potential by appropriate administration of key dietary components leading to early breast cancer prevention in humans. Cancer Prev Res; 11(8); 451-64. ©2018 AACR.

Identification of novel cancer therapeutic targets using a designed and pooled shRNA library screen.

Targeted cancer therapeutics aim to exploit tumor-specific, genetic vulnerabilities specifically affecting neoplastic cells without similarly affecting normal cells. Here we performed sequencing-based screening of an shRNA library on a panel of cancer cells of different origins as well as normal cells. The shRNA library was designed to target a subset of genes previously identified using a whole genome screening approach. This focused shRNA library was infected into cells followed by analysis of enrichment and depletion of the shRNAs over the course of cell proliferation. We developed a bootstrap likelihood ratio test for the interpretation of the effects of multiple shRNAs over multiple cell line passages. Our analysis identified 44 genes whose depletion preferentially inhibited the growth of cancer cells. Among these genes ribosomal protein RPL35A, putative RNA helicase DDX24, and coatomer complex I (COPI) subunit ARCN1 most significantly inhibited growth of multiple cancer cell lines without affecting normal cell growth and survival. Further investigation revealed that the growth inhibition caused by DDX24 depletion is independent of p53 status underlining its value as a drug target. Overall, our study establishes a new approach for the analysis of proliferation-based shRNA selection strategies and identifies new targets for the development of cancer therapeutics.

O-Linked N-Acetylglucosamine (O-GlcNAc) Expression Levels Epigenetically Regulate Colon Cancer Tumorigenesis by Affecting the Cancer Stem Cell Compartment via Modulating Expression of Transcriptional Factor MYBL1.

To study the regulation of colorectal adenocarcinoma progression by O-GlcNAc, we have focused on the O-GlcNAc-mediated epigenetic regulation of human colon cancer stem cells (CCSC). Xenograft tumors from colon tumor cells with O-linked N-acetylglucosamine transferase (OGT) knockdown grew significantly slower than those formed from control cells, indicating a reduced proliferation of tumor cells due to inhibition of OGT expression. Significant reduction of the CCSC population was observed in the tumor cells after OGT knockdown, whereas tumor cells treated with the O-GlcNAcase inhibitor showed an increased CCSC population, indicating that O-GlcNAc levels regulated the CCSC compartment. When grown in suspension, tumor cells with OGT knockdown showed a reduced ability to form tumorspheres, indicating a reduced self-renewal of CCSC due to reduced levels of O-GlcNAc. ChIP-sequencing experiments using an anti-O-GlcNAc antibody revealed significant chromatin enrichment of O-GlcNAc-modified proteins at the promoter of the transcription factor MYBL1, which was also characterized by the presence of H3K27me3. RNA-sequencing analysis showed an increased expression of MYBL1 in tumor cells with OGT knockdown. Forced overexpression of MYBL1 led to a reduced population of CCSC and tumor growth in vivo, similar to the effects of OGT silencing. Moreover, two CpG islands near the transcription start site of MYBL1 were identified, and O-GlcNAc levels regulated their methylation status. These results strongly argue that O-GlcNAc epigenetically regulates MYBL1, functioning similarly to H3K27me3. The aberrant CCSC compartment observed after modulating O-GlcNAc levels is therefore likely to result, at least in part, from the epigenetic regulation of MYBL1 expression by O-GlcNAc, thereby significantly affecting tumor progression.

Identification and characterization of HPV-independent cervical cancers.

BACKGROUND: Human papillomavirus (HPV) initiates cervical cancer, and continuous expression of HPV oncogenes E6 and E7 is thought to be necessary to maintain malignant growth. Current therapies target proliferating cells, rather than specific pathways, and most experimental therapies specifically target E6/E7. We investigated the presence and expression of HPV in cervical cancer, to correlate HPV oncogene expression with clinical and molecular features of these tumors that may be relevant to new targeted therapies. RESULTS: While virtually all cervical cancers contained HPV DNA, and most expressed E6/E7 (HPV-active), a subset (8%) of HPV DNA-positive cervical cancers did not express HPV transcripts (HPV-inactive). HPV-inactive tumors occurred in older women (median 54 vs. 45 years, p = 0.02) and were associated with poorer survival (median 715 vs 3046 days, p = 0.0003). Gene expression profiles of HPV-active and -inactive tumors were distinct. HPV-active tumors expressed E2F target genes and increased AKT/MTOR signaling. HPV-inactive tumors had increased WNT/ß-catenin and Sonic Hedgehog signaling. Substantial genome-wide differences in DNA methylation were observed. HPV-inactive tumors had a global decrease in DNA methylation; however, many promoter-associated CpGs were hypermethylated. Many inflammatory response genes showed promoter methylation and decreased expression. The somatic mutation landscapes were significantly different. HPV-active tumors carried few somatic mutations in driver genes, whereas HPV-inactive tumors were enriched for non-synonymous somatic mutations (p-value < 0.0000001) specifically targeting TP53, ARID, WNT, and PI3K pathways. MATERIALS AND METHODS: The Cancer Genome Atlas (TCGA) cervical cancer data were analyzed. CONCLUSIONS: Many of the gene expression changes and somatic mutations found in HPV-inactive tumors alter pathways for which targeted therapeutics are available. Treatment strategies focused on WNT, PI3K, or TP53 mutations may be effective against HPV-inactive tumors and could improve survival for these cervical cancer patients.

Pathway-Structured Predictive Model for Cancer Survival Prediction: A Two-Stage Approach.

Heterogeneity in terms of tumor characteristics, prognosis, and survival among cancer patients has been a persistent problem for many decades. Currently, prognosis and outcome predictions are made based on clinical factors and/or by incorporating molecular profiling data. However, inaccurate prognosis and prediction may result by using only clinical or molecular information directly. One of the main shortcomings of past studies is the failure to incorporate prior biological information into the predictive model, given strong evidence of the pathway-based genetic nature of cancer, i.e., the potential for oncogenes to be grouped into pathways based on biological functions such as cell survival, proliferation, and metastatic dissemination. To address this problem, we propose a two-stage approach to incorporate pathway information into the prognostic modeling using large-scale gene expression data. In the first stage, we fit all predictors within each pathway using the penalized Cox model and Bayesian hierarchical Cox model. In the second stage, we combine the cross-validated prognostic scores of all pathways obtained in the first stage as new predictors to build an integrated prognostic model for prediction. We apply the proposed method to analyze two independent breast and ovarian cancer datasets from The Cancer Genome Atlas (TCGA), predicting overall survival using large-scale gene expression profiling data. The results from both datasets show that the proposed approach not only improves survival prediction compared with the alternative analyses that ignore the pathway information, but also identifies significant biological pathways.

Association of FGD1 polymorphisms with early-onset breast cancer.

Recent cancer studies have suggested that the faciogenital dysplasia 1 (FGD1) gene may play a role in the development of tumor cells. Somatic alterations in the FGD1 gene and increased Fgd1 protein expression have been observed in many breast tumor cases. The present study sequenced the FGD1 gene in tumor DNA from 46 breast cancer patients using Ion Torrent sequencing. Three synonymous polymorphisms and one missense polymorphism were detected with next-generation sequencing; however, no somatic mutations were observed. The Thr697 variant was identified in 18 patients with an average age at diagnosis of 55 years, which was a lower average age than patients without the polymorphism. In addition, a higher frequency of Thr697 was observed in African-American patients. The Pro712 was observed in 15 breast cancer patients with an average age of 58 years, and was observed as a haplotype with the Thr697 variant in 28% of the breast cancer patients studied. The missense polymorphism (Ala226Thr) was identified in a 40-year-old female patient who had a recurrence of cancer. These polymorphisms (Ala226Thr, Thr697 and Pro712) may be associated with an earlier onset of breast cancer.