An integrative DNA methylation model for improved prognostication of postsurgery recurrence and therapy in prostate cancer patients.

PURPOSE: Patients with clinically localized, high-risk prostate cancer are often treated with surgery, but exhibit variable prognosis requiring long-term monitoring. An ongoing challenge for such patients is developing optimal strategies and biomarkers capable of differentiating between men at risk of early recurrence (<3 years) that will benefit from adjuvant therapies and men at risk of late recurrence (>5 years) who will benefit from long-term monitoring and/or salvage therapies. PATIENTS AND METHODS: DNA methylation changes for 12 genes associated with disease progression were analyzed in 453 prostate tumors. A 4-gene prognostic model (4-G model) for biochemical recurrence (BCR) was derived utilizing LASSO from Cohort 1 (n = 254) and validated in Cohort 2 (n = 199). Subsequently, the 4-G model was evaluated for its association with salvage radiotherapy (RT) and/or hormone therapy, and the additive potential to CAPRA-S to develop an integrative gene model was assessed. RESULTS: The 4-G model was significantly associated with BCR in both cohorts (chi-squared analysis P≤ 0.004) and specifically, with late recurrence at 5+ years (P < 0.001, Cohort 1; P= 0.028, Cohort 2). Multivariable Cox proportional regression analysis identified the 4-G model as significantly associated with salvage RT or hormone therapy in Cohort 1 (hazard ratio (HR) 1.64, 95% confidence interval (CI) 1.29-2.10, P< 0.001) and further validated in Cohort 2 (HR 1.63, 95% CI 1.18-2.25, P< 0.001). The integrative model outperformed prostate-specific antigen and the 4-G model alone for predicting BCR and was associated with patients who received hormone therapy 3+ years postsurgery. CONCLUSIONS: We have identified and validated a novel integrative gene model as an independent prognosticator of BCR and demonstrated its association with late BCR. These patients require more long-term postsurgical monitoring and could be spared the comorbidities of adjuvant therapies.

Temporal Stability and Prognostic Biomarker Potential of the Prostate Cancer Urine miRNA Transcriptome.

BACKGROUND: The development of noninvasive tests for the early detection of aggressive prostate tumors is a major unmet clinical need. miRNAs are promising noninvasive biomarkers: they play essential roles in tumorigenesis, are stable under diverse analytical conditions, and can be detected in body fluids. METHODS: We measured the longitudinal stability of 673 miRNAs by collecting serial urine samples from 10 patients with localized prostate cancer. We then measured temporally stable miRNAs in an independent training cohort (n = 99) and created a biomarker predictive of Gleason grade using machine-learning techniques. Finally, we validated this biomarker in an independent validation cohort (n = 40). RESULTS: We found that each individual has a specific urine miRNA fingerprint. These fingerprints are temporally stable and associated with specific biological functions. We identified seven miRNAs that were stable over time within individual patients and integrated them with machine-learning techniques to create a novel biomarker for prostate cancer that overcomes interindividual variability. Our urine biomarker robustly identified high-risk patients and achieved similar accuracy as tissue-based prognostic markers (area under the receiver operating characteristic = 0.72, 95% confidence interval = 0.69 to 0.76 in the training cohort, and area under the receiver operating characteristic curve = 0.74, 95% confidence interval = 0.55 to 0.92 in the validation cohort). CONCLUSIONS: These data highlight the importance of quantifying intra- and intertumoral heterogeneity in biomarker development. This noninvasive biomarker may usefully supplement invasive or expensive radiologic- and tissue-based assays.

Methylation Markers in Prostate Biopsies Are Prognosticators for Late Biochemical Recurrence and Therapy after Surgery in Prostate Cancer Patients.

After diagnosis of prostate cancer is confirmed by a positive biopsy, the tumor may be surgically removed via radical prostatectomy (RP). However, many prostate cancer patients experience biochemical recurrence after surgery and/or undergo salvage radiotherapy or hormone therapy. Timely treatment is required to prevent the spread of disease in these cases, and biopsy tissue may hold potential for disease prognostication before surgery is ever performed. We previously developed a prognostic multigene methylation panel in RP specimens, including APC, CRIP3, HOXD3, and TGFB2. In the current study, this panel was applied to a cohort of biopsy specimens (n = 86), which were assessed for DNA methylation using the real-time quantitative PCR-based multiplex MethyLight. The biopsy-based methylation panel is significantly associated with biochemical recurrence when combined with the current clinical parameter of prostate-specific antigen (PSA) levels at diagnosis and is able to prognosticate the initiation of salvage radiotherapy, where it outperforms PSA, and/or hormone therapy after RP. In addition, this methylation panel is significantly associated with late recurrence occurring within 5 and 7 years after surgery, when combined with PSA at diagnosis. Combining DNA methylation and clinicopathologic markers at the biopsy stage will not only increase their prognostic ability but will also ensure effective patient management.

GBX2 Methylation Is a Novel Prognostic Biomarker and Improves Prediction of Biochemical Recurrence Among Patients with Prostate Cancer Negative for Intraductal Carcinoma and Cribriform Architecture.

BACKGROUND: Tumor intraductal carcinoma/cribriform architecture (IDC/C) is associated with an unfavorable prognosis and biochemical recurrence (BCR) in prostate cancer (PCa). Up to 70% of PCa patients are IDC/C-negative, but it is estimated that 20% of these cases still experience BCR. Thus, biomarkers for better detection of aggressive disease in IDC/C-negative patients are required. OBJECTIVE: To investigate tumor-specific methylation of the transcription factor GBX2 as a novel prognosticator and predictor of BCR in PCa patients stratified by histopathologic features including IDC/C. DESIGN, SETTING, AND PARTICIPANTS: Using genome-wide methylome profiling, we identified higher GBX2 methylation in grade group (GG) 4 tumors compared to GG1 (discovery cohort). The prognostic nature of GBX2 methylation was validated in silico using The Cancer Genome Atlas data (n=478) and a quantitative methylation assay for radical prostatectomy samples (n=254). Regulation of GBX2 methylation was investigated in prostate cells using methyl-CpG-binding domain sequencing and methylation analysis in functional knockouts of TET2, a key epigenetic player in prostate carcinogenesis. OUTCOME MEASUREMENTS AND STATISTICAL ANALYSIS: The association of GBX2 methylation with Gleason score (GS), pathologic stage (pT), IDC/C, and BCR was analyzed using Kruskal-Wallis and Mann-Whitney tests. Univariate and multivariate Cox regression analyses were used to predict BCR. RESULTS: GBX2 methylation was associated with GS (p<0.05), pT (p<0.01), and BCR (p<0.05). GBX2 methylation (p=0.004), GS (p<0.001), pT (p=0.012), and prostate-specific antigen (p=0.005) were independent predictors of BCR. Among IDC/C-negative patients, GBX2 methylation improved prediction of BCR (p=0.002). Loss of TET2 in prostate cells resulted in greater GBX2 methylation. CONCLUSIONS: We identified GBX2 methylation as a novel prognostic factor in PCa and an independent predictor of BCR. We demonstrated the additive value of GBX2 methylation in predicting BCR among IDC/C-negative patients and elucidated a novel TET2-mediated upstream epigenetic regulatory mechanism of GBX2. PATIENT SUMMARY: We identified GBX2 methylation as a promising prognostic biomarker that could improve the identification of prostate cancer patients at higher risk of biochemical recurrence.

GLP-1 receptor agonism ameliorates hepatic VLDL overproduction and de novo lipogenesis in insulin resistance.

BACKGROUND/OBJECTIVES: Fasting dyslipidemia is commonly observed in insulin resistant states and mechanistically linked to hepatic overproduction of very low density lipoprotein (VLDL). Recently, the incretin hormone glucagon-like peptide-1 (GLP-1) has been implicated in ameliorating dyslipidemia associated with insulin resistance and reducing hepatic lipid stores. Given that hepatic VLDL production is a key determinant of circulating lipid levels, we investigated the role of both peripheral and central GLP-1 receptor (GLP-1R) agonism in regulation of VLDL production. METHODS: The fructose-fed Syrian golden hamster was employed as a model of diet-induced insulin resistance and VLDL overproduction. Hamsters were treated with the GLP-1R agonist exendin-4 by intraperitoneal (ip) injection for peripheral studies or by intracerebroventricular (ICV) administration into the 3rd ventricle for central studies. Peripheral studies were repeated in vagotomised hamsters. RESULTS: Short term (7-10 day) peripheral exendin-4 enhanced satiety and also prevented fructose-induced fasting dyslipidemia and hyperinsulinemia. These changes were accompanied by decreased fasting plasma glucose levels, reduced hepatic lipid content and decreased levels of VLDL-TG and -apoB100 in plasma. The observed changes in fasting dyslipidemia could be partially explained by reduced respiratory exchange ratio (RER) thereby indicating a switch in energy utilization from carbohydrate to lipid. Additionally, exendin-4 reduced mRNA markers associated with hepatic de novo lipogenesis and inflammation. Despite these observations, GLP-1R activity could not be detected in primary hamster hepatocytes, thus leading to the investigation of a potential brain-liver axis functioning to regulate lipid metabolism. Short term (4 day) central administration of exendin-4 decreased body weight and food consumption and further prevented fructose-induced hypertriglyceridemia. Additionally, the peripheral lipid-lowering effects of exendin-4 were negated in vagotomised hamsters implicating the involvement of parasympathetic signaling. CONCLUSION: Exendin-4 prevents fructose-induced dyslipidemia and hepatic VLDL overproduction in insulin resistance through an indirect mechanism involving altered energy utilization, decreased hepatic lipid synthesis and also requires an intact parasympathetic signaling pathway.