Summary Somatic mutations in IDH1/2 and TET2 result in impaired TET2 mediated conversion of 5-methylcytosine (5-mC) to 5-hydroxymethylcytosine (5-hmC). The observation that WT1 inactivating mutations anti-correlate with TET2/IDH1/2 mutations in AML led us to hypothesize that WT1 mutations may impact TET2 function. WT1 mutant acute myeloid leukemia (AML) patients have reduced 5-hmC levels similar to TET2/IDH1/2-mutant AML. These mutations are characterized by convergent, site-specific alterations in DNA hydroxymethylation, which drive differential gene expression more than alterations in DNA promoter methylation. WT1 overexpression increases global levels of 5-hmC, and WT1 silencing reduced 5-hmC levels. WT1 physically interacts with TET2 and TET3, and WT1 loss of function results in a similar hematopoietic differentiation phenotype as observed with TET2 deficiency. These data provide a novel role for WT1 in regulating DNA hydroxymethylation and suggest that TET2 IDH1/2, and WT1 mutations define a novel AML subtype defined by dysregulated DNA hydroxymethylation.

The global burden of kidney disease is increasing, and several etiologies first begin in childhood. Risk factors for pediatric kidney disease are common in Africa, but data regarding its prevalence are lacking. We completed a systematic review of community-based studies describing the prevalence of proteinuria, hematuria, abnormal imaging, or kidney dysfunction among children in sub-Saharan Africa. Medline and Embase were searched. Five hundred twenty-three references were reviewed. Thirty-two references from 9 countries in sub-Saharan Africa were included in the qualitative synthesis. The degree of kidney damage and abnormal imaging varied widely: proteinuria 32.5% (2.2%-56.0%); hematuria 31.1% (0.6%-67.0%); hydronephrosis 11.3% (0.0%-38.0%), hydroureter 7.5% (0.0%-26.4%), major kidney abnormalities 0.1% (0.0%-0.8%). Serum creatinine was reported in four studies with insufficient detail to identify the prevalence renal dysfunction. A majority of the studies were performed in Schistosoma haematobium endemic areas. A lower prevalence of kidney disease was observed in the few studies from non-endemic areas. Published data on pediatric kidney disease in sub-Saharan Africa is highly variable and dependent on S. haematobium prevalence. More community-based studies are needed to describe the burden of pediatric kidney disease, particularly in regions where S. haematobium infection is non-endemic.

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BACKGROUND A positive surgical margin (PSM) following radical prostatectomy (RP) for prostate cancer (PCa) is associated with increased risk of biochemical recurrence. We sought to examine whether the pathologist is an independent predictor of PSMs. METHODS We performed a retrospective review of 3,557 men who underwent RP for localized PCa at our institution from 2003–2015. We evaluated 29 separate pathologists. Univariate and multivariable logistic regression were used to test variables previously shown to influence PSM rates. RESULTS Overall rate of PSM was 18.9%. Compared to patients without PSM, patients with PSM had higher BMI (mean 28.8 vs. 28.3), Gleason ≥ 7 (84% vs. 66%), extracapsular extension (51% vs. 20%), and median PSA (5.9 vs. 5.1 ng/ml) (all p < 0.05). Univariate logistic regression showed surgeon experience, pathologist experience, and pathologist GU fellowship training were all predictors of PSMs (all p < 0.05). Multivariable regression analysis confirmed decreased surgeon experience, increased pathologist experience, higher pathologic Gleason score, higher pathologic stage, and higher PSA were significant predictors of PSMs. Increasing surgeon experience was associated with decreased odds of PSM (OR 0.79 per 1 SD increase, 95% CI [0.70 – 0.89]). In contrast, increasing pathologist experience was associated with increased odds of PSM (OR 1.11 per 1 SD increase, 95% CI [1.03 – 1.19]). The relationship between pathologist experience and PSM appeared to be non-linear (Figure 2). CONCLUSIONS Greater pathologist experience appears to be associated with greater odds of PSMs following radical prostatectomy, even after controlling for case mix, pathologist fellowship training, and surgeon experience. Based on these findings, pathologists with less experience reviewing RP specimens may consider requesting re-review by a dedicated GU pathologist.

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Purpose:Adrenocortical carcinoma is a rare but aggressive malignancy. While centralization of care to referral centers improves outcomes across common urological malignancies, there exists a paucity of data for low-incidence cancers. We sought to evaluate differences in practice patterns and overall survival in patients with adrenocortical carcinoma across types of treating facilities.Materials and Methods:We identified all patients diagnosed with adrenocortical carcinoma from 2004–2016 in the National Cancer Database. The Kaplan-Meier method was used to evaluate overall survival and multivariable Cox regression analysis was used to investigate independent predictors of overall survival. The chi-square test was used to analyze differences in practice patterns.Results:We identified 2,886 patients with adrenocortical carcinoma. Median overall survival was 21.8 months (95% CI 19.8–23.8). Academic centers had improved overall survival versus community centers on unadjusted Kaplan-Meier analysis (p <0.05) and had higher rates of adrenalectomy or radical en bloc resection (p <0.001), performed more open surgery (p <0.001), administered more systemic therapy (p <0.001) and had lower rates of positive surgical margins (p=0.03). On multivariable analysis, controlling for treatment modality, academic centers were associated with significantly decreased risk of death (HR 0.779, 95% CI 0.631–0.963, p=0.021).Conclusions:Treatment of adrenocortical carcinoma at an academic center is associated with improved overall survival compared to community programs. There are significant differences in practice patterns, including more aggressive surgical treatment at academic facilities, but the survival benefit persists on multivariable analysis controlling for treatment modality. Further studies are needed to identify the most important predictors of survival in this at-risk population.

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Mutations in TET2 and IDH1/2, which result in reduced conversion of 5-methylcytosine (5-mC) to 5-hydroxymethylcytosine (5-hmC), are observed in a significant proportion of patients with acute myeloid leukemia (AML). Hence these mutations define a novel class of AMLs with alterations in DNA methylation. However, it has been shown that not all AML cases with low levels of 5-hmC harbor somatic mutations in TET2 and IDH1/2. We hypothesized there are additional somatic mutations that can lead to alterations in TET enzyme function and resultant changes in the epigenetic state of hematopoietic cells. Mutational studies of the ECOG 1900 trial allowed us to discover that somatic mutations in the WT1 gene were mutually exclusive of IDH1/2 mutations and negatively correlated with TET2 mutations (Figure 1A). 28/313 (9%) of TET2/IDH-wild-type patients had somatic WT1 mutations, whereas 2/85 (2%) TET2/IDH1/2-mutant patients had co-occurring WT1 mutations (p=0.026, Fisher's Exact test). Analysis of combined data from the ECOG 1900 study and the AML TCGA dataset confirmed a significant anti-correlation between WT1 mutations and TET2/IDH1/2 mutations (p=0.016). We next compared the DNA methylation profiles of WT1, TET2, and IDH1/2 mutant AML patients to AML patients wild-type for these disease alleles. Comparative analysis of the three hypermethylation profiles revealed a near-complete overlap of TET2 and WT1 hypermethylated loci within the IDH1/2 hypermethylation signature, and a highly significant overlap between the TET2 and WT1 mutant signatures (p-value< 0.001 for all comparisons), consistent with convergent, site-specific effects on DNA methylation. These genetic observations suggested a shared functional role for WT1, TET2, and IDH1/2 mutations in AML in regulating 5-hmC. WT1-mutant AML patients had significantly reduced 5-hmC (Liquid chromatography-electron spray ionization-tandem mass spectrometry) compared to AML patients wild-type for WT1, TET2 or IDH1/2 (p=0.016 T-test Figure 1B), similar to the reduction in 5-hmC observed in IDH1/2-mutant and TET2-mutant AML patient samples. In order to determine the impact of IDH1/2, WT1 and TET2 mutations on the distribution of epigenetic marks throughout the genome more precisely, we examined 5-hmC localization in primary AML specimens with next-generation sequencing. The average number of 5-hmC peaks present in samples from AML patients with TET2, WT1, IDH1 or IDH2 mutations was significantly reduced compared to AML patients wild-type for these disease alleles (t-test p-values between 0.0005 and 0.003). We observed highly significant overlap of differential 5-hmC peaks lost in WT1-mutant AML and TET2-mutant AML (Hypergeometric test p-value < 10-133). In a manner analogous to the findings for DNA methylation, the regions with reduced 5-hmC in WT1/TET2 mutant AML were a subset of those found in IDH1/2 mutant AML patients. We next investigated whether perturbations in WT1 expression can directly alter 5-hmC levels. Knockdown of Wt1 significantly decreased 5-hmC levels in M15 cells (p<0.01) (Figure 1C). Conversely, overexpression of WT1 in 32D cells significantly increased 5-hmC levels compared to cells expressing a control vector or WT1-mutant (p<0.05) (Figure 1D). Thus, WT1 can directly regulate 5-hmC levels. Given the effects of WT1 on 5-hmC levels and the inverse correlation between WT1 and TET2 mutations in AML, we hypothesized that WT1 might modulate TET2 function through direct interaction.Co-immunoprecipitation experiments demonstrated WT1 interacts with TET2 and with TET3. Consistent with these effects, WT1 expression inhibited colony forming ability of TET2-deficient cells, but could not attenuate the colony growth of TET2/TET3-deficient cells in vitro. Taken together, our genetic and functional studies demonstrate an inverse correlation between WT1 mutations and TET2/IDH1/2 mutations in AML and a functional role for WT1 in regulating DNA methylation. WT1 mutant AML is characterized by similar global and site-specific alterations in 5-hmC/5-mC as observed in TET2/IDH1/2 mutant AML. Furthermore, we demonstrate that alterations in WT1 levels directly regulate 5-hmC levels, which is due to an interaction between TET2/TET3 and WT1. These data provide a novel role for WT1 in regulating DNA hydroxymethylation and suggest that TET2 IDH1/2, and WT1 mutations define a novel AML subtype. Figure 1 Figure 1. Disclosures Levine: Novartis: Consultancy, Grant support Other.