Reactivation Viremia in Pediatric Sepsis.

OBJECTIVES: Reactivation viremia is associated with adverse clinical outcomes and immune dysfunction in adults with sepsis. We determined the incidence of viremia and its association with clinical outcomes and immune paralysis phenotype in children with severe sepsis. DESIGN: Prospective cohort study. SETTING: Single academic PICU from September 2016 to March 2018. PATIENTS: Fifty-nine patients 2-17 years old treated for severe sepsis. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: We performed real-time polymerase chain reaction assays on whole blood specimens to determine the incidence of cytomegalovirus. Cytomegalovirus was detected in three patients (5%). All patients with cytomegalovirus viremia were seropositive, with an incidence of 13% in this subset. We additionally performed Epstein-Barr virus and human herpesvirus-6 polymerase chain reaction assays on last available specimens and detected Epstein-Barr virus in 4% and human herpesvirus-6 in 30% of the study population. Overall, viremia was not associated with clinical outcomes or immune function in univariable analyses. However, viremia was associated with lower odds of complicated course (defined as death within 28 d or ≥ 2 organ dysfunctions at 7 d) after controlling for age, Pediatric Risk of Mortality III score, and blood transfusion (adjusted odds ratio, 0.08; 95% CI, 0.01-0.84; p = 0.04). CONCLUSIONS: Children with severe sepsis had low rates of detectable viremia, which limited analyses of its association with clinical outcomes or immune paralysis phenotype. Given the rare occurrence of cytomegalovirus viremia, in particular, our study does not support a role for viremia as a biomarker of illness severity or as a modifiable risk factor of clinical outcomes for most patients. Future studies on the role of viremia in pediatric sepsis will need to consider the challenges posed by low rates of viremia in this population.

Association of Delayed Antimicrobial Therapy with One-Year Mortality in Pediatric Sepsis.

OBJECTIVE: Delayed antimicrobial therapy in sepsis is associated with increased hospital mortality, but the impact of antimicrobial timing on long-term outcomes is unknown. We tested the hypothesis that hourly delays to antimicrobial therapy are associated with 1-year mortality in pediatric severe sepsis. DESIGN: Retrospective observational study. SETTING: Quaternary academic pediatric intensive care unit (PICU) from February 1, 2012 to June 30, 2013. PATIENTS: One hundred sixty patients aged ≤21 years treated for severe sepsis. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: We tested the association of hourly delays from sepsis recognition to antimicrobial administration with 1-year mortality using multivariable Cox and logistic regression. Overall 1-year mortality was 24% (39 patients), of whom 46% died after index PICU discharge. Median time from sepsis recognition to antimicrobial therapy was 137 min (IQR 65-287). After adjusting for severity of illness and comorbid conditions, hourly delays up to 3 h were not associated with 1-year mortality. However, increased 1-year mortality was evident in patients who received antimicrobials ≤1 h (aOR 3.8, 95% CI 1.2, 11.7) or >3 h (aOR 3.5, 95% CI 1.3, 9.8) compared with patients who received antimicrobials within 1 to 3 h from sepsis recognition. For the subset of patients who survived index PICU admission, antimicrobial therapy ≤1 h was also associated with increased 1-year mortality (aOR 5.5, 95% CI 1.1, 27.4), while antimicrobial therapy >3 h was not associated with 1-year mortality (aOR 2.2, 95% CI 0.5, 11.0). CONCLUSIONS: Hourly delays to antimicrobial therapy, up to 3 h, were not associated with 1-year mortality in pediatric severe sepsis in this study. The finding that antimicrobial therapy ≤1 h from sepsis recognition was associated with increased 1-year mortality should be regarded as hypothesis-generating for future studies.

Inactivation of the tumor suppressor WTX in a subset of pediatric tumors.

WTX is a tumor suppressor gene expressed during embryonic development and inactivated in 20-30% of cases of Wilms tumor, the most common pediatric kidney cancer. WTX has been implicated in several cellular processes including Wnt signaling, WT1 transcription, NRF2 degradation, and p53 function. Given that WTX is widely expressed during embryonic development and has been recently shown to regulate mesenchymal precursor cells in several organs, we tested for the potential involvement of WTX in a panel of pediatric tumors and adult sarcomas. A total of 353 tumors were screened for WTX deletions by fluorescence in situ hybridization (FISH). Discrete somatic WTX deletions were identified in two cases, one hepatoblastoma and one embryonal rhabdomyosarcoma, and confirmed by array comparative genomic hybridization. Direct sequencing of the full WTX open reading frame in 24 hepatoblastomas and 21 embryonal rhabdomyosarcomas did not identify additional mutations in these tumor types. The presence of WTX mRNA was confirmed in hepatoblastomas and embryonal rhabdomyosarcomas without WTX deletions by RNA-in situ hybridization. Notably, tumors with evidence of WTX inactivation, Wilms tumor, hepatoblastoma and rhabdomyosarcoma, are primitive tumors that resemble undifferentiated precursor cells and are linked to overgrowth syndromes. These results indicate that WTX inactivation occurs in a wider variety of tumor types than previously appreciated and point to shared pathogenic mechanisms between a subset of pediatric malignancies.

Wilms' tumor with an apparently balanced translocation t(X;18) resulting in deletion of the WTX gene.

The recent description of a new X chromosome tumor suppressor gene, WTX, that is commonly inactivated in Wilms' tumor prompted us to examine the possible involvement of WTX in a case of Wilms' tumor containing an apparently balanced reciprocal translocation between chromosomes X and 18 (t(X;18)(q11;p11)). Fluorescence in situ hybridization (FISH) analysis of paraffin tumor sections indeed revealed a deletion of the WTX locus at Xq11. High-resolution array comparative genomic hybridization (array CGH) analysis of tumor DNA revealed a 1.5 Mb chromosome deletion encompassing the WTX gene at Xq11. No loss of genetic material was detected on chromosome 18. Interestingly, unlike most tumors with acquired chromosomal translocations, where a new fusion oncogene or promoter-oncogene fusion is created and drives tumor growth, the t(X;18) in this tumor appears to drive tumorigenesis via deletion of a tumor suppressor. This case demonstrates the importance of array CGH and FISH as adjuncts in tumor cytogenetics and in identifying pathogenic microdeletions in "balanced" translocations that are not truly balanced.

Mucinous differentiation correlates with absence of EGFR mutation and presence of KRAS mutation in lung adenocarcinomas with bronchioloalveolar features.

Somatic mutations in the epidermal growth factor receptor gene (EGFR) are detected in a subset of lung adenocarcinomas, particularly bronchioloalveolar carcinoma (BAC) and adenocarcinoma with bronchioloalveolar features (AWBF), and correlate with clinical response to tyrosine kinase inhibitors (TKIs). In contrast, lung adenocarcinomas refractory to TKIs often have activating mutations in KRAS but lack EGFR mutations. Some adenocarcinomas have mucinous histology, but the clinical and molecular significance of the mucinous pattern is less well studied. We analyzed 43 BAC and AWBF tumors submitted for EGFR mutation testing to identify histopathological features that predicted EGFR or KRAS mutations. EGFR mutations were detected in 14 of 30 (47%) nonmucinous tumors, whereas 0 of 13 mucinous tumors harbored an EGFR mutation (P = 0.003). Missense mutations in KRAS codon 12 were detected in six of seven (86%) mucinous adenocarcinomas but only 3 of 18 (17%) nonmucinous adenocarcinomas (P = 0.003). Thus, in BAC/AWBF mucinous differentiation was significantly correlated with the absence of EGFR mutation and presence of KRAS mutation, suggesting that mucinous BACs/AWBFs are unlikely to respond to TKIs. Therefore, our data suggest that EGFR sequence analysis could be avoided in BAC/AWBF when true mucinous morphology is identified, avoiding the associated testing costs.

An X chromosome gene, WTX, is commonly inactivated in Wilms tumor.

Wilms tumor is a pediatric kidney cancer associated with inactivation of the WT1 tumor-suppressor gene in 5 to 10% of cases. Using a high-resolution screen for DNA copy-number alterations in Wilms tumor, we identified somatic deletions targeting a previously uncharacterized gene on the X chromosome. This gene, which we call WTX, is inactivated in approximately one-third of Wilms tumors (15 of 51 tumors). Tumors with mutations in WTX lack WT1 mutations, and both genes share a restricted temporal and spatial expression pattern in normal renal precursors. In contrast to biallelic inactivation of autosomal tumor-suppressor genes, WTX is inactivated by a monoallelic "single-hit" event targeting the single X chromosome in tumors from males and the active X chromosome in tumors from females.

Expression of oligodendroglial and astrocytic lineage markers in diffuse gliomas: use of YKL-40, ApoE, ASCL1, and NKX2-2.

The phenotypic heterogeneity of astrocytic and oligodendroglial tumor cells complicates establishing accurate diagnostic criteria, and lineage-specific markers would facilitate diagnosis of glioma subtypes. Based on data from the literature and from expression microarrays, we selected molecules relevant to gliogenesis and glial lineage specificity and then used immunohistochemistry to assess expression of these molecules in 55 diffuse gliomas, including 8 biphasic oligoastrocytomas, 21 oligodendrogliomas (all with 1p/19qloss), 21 astrocytomas, and 5 glioblastomas. For the astrocytic lineage markers (GFAP, YKL-40, and ApoE), GFAP expression was significantly higher in the astrocytic component of oligoastrocytomas compared with the oligodendroglial part; similar patterns were detected for YKL-40 and ApoE, although the differences were not significant. GFAP, YKL-40, and ApoE reliably distinguished grade II-III oligodendrogliomas from grade II-IV astrocytomas (p < 0.0001, p = 0.002, and p < 0.0001, respectively). Among the oligodendroglial lineage markers (Olig2, Sox10, ASCL1, and NKX2-2), ASCL1 and NKX2-2 displayed significantly different immunostaining between oligodendrogliomas and astrocytomas (p = 0.017 and 0.004, respectively), but none clearly differentiated between the 2 glial populations of oligoastrocytomas. In addition to GFAP, therefore, YKL-40, ApoE, ASCL1, and NKX2-2 represent promising tumor cell markers to distinguish oligodendrogliomas from astrocytomas.