Development of a Clinically Applicable NanoString-Based Gene Expression Classifier for Muscle-Invasive Bladder Cancer Molecular Stratification.

Transcriptional profiling of muscle-invasive bladder cancer (MIBC) using RNA sequencing (RNA-seq) technology has demonstrated the existence of intrinsic basal and luminal molecular subtypes that vary in their prognosis and response to therapy. However, routine use of RNA-seq in a clinical setting is restricted by cost and technical difficulties. Herein, we provide a single-sample NanoString-based seven-gene (KRT5, KRT6C, SERPINB13, UPK1A, UPK2, UPK3A and KRT20) MIBC molecular classifier that assigns a luminal and basal molecular subtype. The classifier was developed in a series of 138 chemotherapy naïve MIBCs split into training (70%) and testing (30%) datasets. Further, we validated the previously published CK5/6 and GATA3 immunohistochemical classifier which showed high concordance of 96.9% with the NanoString-based gene expression classifier. Immunohistochemistry-based molecular subtypes significantly correlated with recurrence-free survival (RFS) and disease-specific survival (DSS) in univariable (p = 0.006 and p = 0.011, respectively) and multivariate cox regression analysis for DSS (p = 0.032). Used sequentially, the immunohistochemical- and NanoString-based classifiers provide faster turnaround time, lower cost per sample and simpler data analysis for ease of clinical implementation in routine diagnostics.

D2-40 is expressed on the luminal surface of pulmonary airspaces in normal developing and adult lung but is lost in conditions associated with intra-alveolar infiltrates.

The D2-40 antigen is a glycosylated sialomucin that is strongly expressed by lymphatic endothelial cells. Recently we observed the expression of D2-40 on the luminal surface of pulmonary airspaces in lung sections. The aim of the study was to assess the expression of D2-40 antigen in normal lung development and in various pathologic conditions in which abnormal alveolar infiltrates were present. Formalin-fixed lung tissue was selected from 42 fetal/neonatal autopsy cases ranging in gestational age from 12 to 41 weeks and from 10 adult lungs. In the fetal/neonatal group, 22 cases were histologically normal, whereas 20 were abnormal (including cases of pneumonia, alveolar hemorrhage, meconium aspiration, pulmonary hypoplasia, and pulmonary interstitial emphysema). In the adult group, 5 cases were histologically normal and 5 had pneumonia. Immunohistochemical staining was performed on all cases using antibody to D2-40. All cases of normal fetal/neonatal lung and normal adult lung showed diffuse strong expression of D2-40 on the luminal surface of the alveolar lining cells. D2-40 expression was also noted on the bronchiolar lining cells of normal fetal/neonatal lung. In all cases in which there was an abnormal infiltrate or foreign material within the airspaces, expression of D2-40 was lost in the alveolar lining. The production of the D2-40 antigen in the alveolar lining occurs as early as 12 weeks gestation and continues to be present throughout all other stages of lung development, as well as in adult lung. These results suggest that D2-40 may have a cell membrane protective function.

Significance of p53 expression in immature teratomas.

Twenty-nine pediatric immature teratomas were reviewed to determine the frequency and clinical significance of p53 expression. Tumors were stained for p53 expression by immunohistochemistry and results were correlated with the presence of other germ cell tumor elements and with outcome. Sequencing of p53 for mutations was performed on positive cases. Eighteen cases showed widespread positive p53 staining of the immature teratoma elements, 9 showed staining only in very occasional cells, and 2 cases showed no staining. Of the 18 positive cases, 5 recurred. All five were pure immature teratomas at diagnosis. Four recurred as immature or mature teratoma and one as a sarcoma; all except one showed frequent cells positive for p53 in the recurrent tumor. Another 5 of the 18 diffusely positive cases contained immature teratoma as well as other malignant germ cell elements at diagnosis; none of these recurred. None of the remaining eight cases with frequent positive cells, the nine cases with occasionally positive p53 staining, or the two cases with no staining recurred or demonstrated other germ cell tumor elements. We conclude that p53 expression is not unusual in immature teratoma and diffuse p53 immunopositivity is associated with recurrence or the presence of malignant elements in approximately 50% of cases. In only 1 of 29 cases tested was p53 immunopositivity associated with mutations in the p53 gene; hence, overexpression in the majority of cases is presumed to reflect increased half-life of the protein from undetermined stabilizing factors. Expression of p21, a p53 target gene, was only focal, suggesting impaired transcriptional activation by p53. The finding of frequent p53-positive cells in immature teratoma should prompt a search for malignant elements within the tumor and affected patients should be followed closely for evidence of recurrence.

Fulminant disseminated Varicella Zoster virus infection without skin involvement.

BACKGROUND: Varicella Zoster virus (VZV) infection is potentially very serious in bone marrow transplant recipients, and may manifest as a disseminated visceral infection. This condition is generally accompanied by a vesicular rash. OBJECTIVES: We review here a case of fulminant fatal disseminated VZV infection, not accompanied by skin involvement, and the laboratory approaches currently available to diagnose this disease. STUDY DESIGN: Post mortem tissue samples were subjected to histopathological examination, and tested for herpesviruses by electron microscopy and PCR. RESULTS: Intranuclear inclusions were noted by histological examination in the lungs, liver, kidneys and bone marrow. Particles with a herpesvirus morphology were visualized in liver tissue. VZV DNA was detected in liver and bone marrow by PCR followed by sequencing of the amplicons. Viremia was documented by retrospective testing of the serum by PCR. CONCLUSIONS: A disseminated VZV infection which proved rapidly fatal was demonstrated in a case without skin manifestations. This rare presentation of VZV infection is potentially underdiagnosed. Testing for VZV viremia by PCR can at the very least suggest the diagnosis although whether plasma-associated viremia is truly pathognomonic of visceral disseminated infection remains to be established.