Deborah A. Belchis

Inflammatory cell types and clinical features of interstitial cystitis.

PURPOSE We tested whether the types of inflammatory cells seen on bladder biopsies were associated with other clinical features and urinary markers of interstitial cystitis. MATERIALS AND METHODS Bladder biopsies from 30 interstitial cystitis patients were evaluated by immunohistochemical staining for T cells, B cells, macrophages and human leukocyte antigen-DR positive cells. These findings were tested for associations with clinical features and urinary markers of interstitial cystitis using alpha = 0.01 because multiple tests were performed. RESULTS Overall severity of inflammation was significantly associated with age at symptom onset, symptom relief after bladder distention and urinary interleukin-6 levels. Patients with severe inflammation had trends toward smaller bladder capacity under anesthesia, increased bladder vascularity and mucosal cracks, lower urinary MUC-1 glycoprotein levels and absence of bloating as a symptom. B cell staining was significantly associated with severe inflammation, symptom relief after distention and absence of bloating as a symptom. T cell staining was significantly associated with severe inflammation and age at symptom onset. Human leukocyte antigen-DR staining had trends with symptoms, including presence of bloating, constant urge to void and absence of burning. Macrophage staining did not associate with any features tested at the alpha = 0.05 level. CONCLUSIONS Interstitial cystitis patients with severe inflammation have different age, treatment response and urinary marker levels than those with mild inflammation. These findings suggest that the 2 patient groups have different underlying pathophysiologies. The significant associations for T and B cell staining were similar to those for overall inflammation.

Congenital disseminated malignant rhabdoid tumor: a distinct clinicopathologic entity demonstrating abnormalities of chromosome 22q11.

The clinical, pathologic, and immunohistochemical features of a widely disseminated tumor with rhabdoid phenotype are described in nine infants < or = 3 months of age. Five neonates had tumor evident at birth, two of which had placental metastases. The average survival following diagnosis was < 6 weeks. None of the infants had an apparent primary tumor in either the kidney or brain. In four cases, the dominant mass involved the head and neck region, and in two cases, the primary mass was paraspinal. The histologic features were those of a high-grade, round cell neoplasm with abundant cytoplasm and containing cells with cytoplasmic filamentous inclusions. Immunohistochemical studies revealed polyphenotypic antigen expression. Genetic information was available from eight of nine cases. Karyotype analysis revealed abnormalities of chromosome band 22q11-12 in three of six tumors. Fluorescence in situ hybridization studies or molecular studies demonstrated 22q11.2 deletions in all five cases with available frozen tissue, two of which had translocations involving 22q by karyotype analysis. The similar clinical and pathologic findings in these rapidly fatal tumors in infants and the demonstration of abnormalities of chromosome 22q11 in a majority of the cases supports their histogenetic and nosologic relationship to the family of malignant rhabdoid tumors that typically occur in young children in several anatomic sites, including kidney, soft tissues, liver, and brain. Like neuroblastoma and rhabdomyosarcoma, malignant rhabdoid tumor can appear as disseminated disease at birth or shortly thereafter.

Loss of heterozygosity and microsatellite instability at the retinoblastoma locus in osteosarcomas.

Studies of osteosarcoma cell lines or frozen tissue have detected loss of heterozygosity (LOH) at the retinoblastoma (RB) locus by Southern blot analysis or restriction fragment length polymorphism. Most archived clinical specimens cannot be analyzed by these techniques. We analyzed formalin-fixed, paraffin-embedded samples from 19 cases of osteosarcoma for molecular changes at the RB locus using polymerase chain reaction amplification of polymorphic short tandem repeat sequences (microsatellite repeats). Four repeat sequences, two within and two flanking the RB gene, were analyzed. Fourteen of 18 informative cases (78%) showed molecular changes at the RB locus. LOH was identified in 13 cases (72%). Unexpectedly, microsatellite instability (MI) was found in eight cases (44%). All of the cases of MI involved alterations of more than one repeat unit, and six of eight were associated with LOH. LOH was identified at three unlinked loci in one case and at a single locus in another. Microsatellite analysis of archival tissue yields prevalence rates of LOH comparable to those found by other methods and has the added advantage of showing MI. The ability to use formalin-fixed, paraffin-embedded tissue extends genetic analysis to routinely processed surgical material and may permit molecular confirmation of challenging cases of osteosarcoma.

Association of a mosaic chromosomal 22q 11 deletion with hypoplastic left heart syndrome

The atypical presentation of CATCH 22 raises several important concerns. First, in this patient, as in others, the heart defects were found in association with subtle facial abnormalities but with few of the other criteria normally seen in CATCH 22. This association alone may be sufficient to raise suspicion that an interstitial 22q11 deletion may be present. Second, the incidence of chromosome 22 deletions in parents of children with a 22q11 deletion (25%) suggests that siblings or subsequent fetuses may also be at risk. Parents with subtle or unusual manifestations of CATCH 22 may be unaware of their potential carrier status. Finally, the recognition of chromosomal mosaicism in this patient may have been fortuitous, as cytogenetic studies of leukocytes from other individuals with a mosaic karyotype may sometimes fail to reveal a 22q11 deletion that is present in cardiac tissues. Molecular cytogenetic analysis of cardiac specimens that are removed during routine surgical procedures may be warranted in appropriate clinical situations.

Relationships between bladderinflammation and other clinical features in interstitial cystitis

OBJECTIVES Interstitial cystitis (IC) has been considered possibly to represent more than one disease process. If so, patients would be expected to form distinct subgroups. The degree of mononuclear inflammation on bladder biopsy can be objectively quantified and might be a useful parameter for subgroup division. The hypothesis of this study was that patients with mild versus severe inflammation would differ with regard to other clinical features of IC. METHODS Sixteen patients who met the original National Institute of Diabetes, Digestive and Kidney Diseases criteria for IC underwent cystoscopy with bladder distention and biopsy. The degree of mononuclear inflammation on bladder biopsy was classified as mild, with less than 100 mononuclear cells/high power field (HPF), or severe (100 or more mononuclear cells/HPF or lymphoid aggregates). Associations were sought between degree of inflammation and other subjective and objective clinical features. RESULTS Five patients had severe inflammation and 11 had mild inflammation. The major finding was that the patients with severe inflammation experienced better symptom relief after cystoscopy with bladder distention under anesthesia. This difference was highly significant (Fishers exact test, p = 0.0014). For the other clinical features studied, these two groups did not differ significantly. CONCLUSIONS Two distinct IC patient groups were identified by bladder biopsy findings. These two groups had significantly different treatment responses. If this difference is confirmed with a larger number of patients, it would suggest that these two patient groups may have different underlying disease processes.

A Unique, Histopathologic Lesion in a Subset of Patients With Spontaneous Pneumothorax

CONTEXT Spontaneous pneumothorax can be idiopathic (primary), or it can occur in association with an underlying predisposing condition (secondary). Spontaneous pneumothorax may be a harbinger of an undiagnosed clinical condition, which may be associated with serious systemic abnormalities, making early recognition and diagnosis important. The pulmonary pathology of some of these disorders has not been fully elucidated. OBJECTIVE To review cases of pneumothorax in the hope of identifying pathologic features that might correlate to specific clinical syndromes. DESIGN The pathology computer files at 3 hospitals were searched for all cases of spontaneous pneumothorax, primary and secondary, regardless of etiology during a 11-year period. Ninety-two cases were retrieved. Each of the cases was evaluated for reactive eosinophilic pleuritis, elastosis, pleural fibrosis, emphysema, intra-alveolar macrophages, cholesterol clefts, vasculopathy, and intraparenchymal or intrapleural cysts. Clinical information regarding asthma and smoking history, site of the pneumothorax, family history, radiographic findings, predisposing conditions, recurrence, age, and sex were extracted from the medical records. RESULTS In 11 patients (12% of all the patients with spontaneous pneumothorax), a distinctive pattern of pleural fibrosis with islands of fibroblastic foci within a myxoid stroma was noted at the pleural-parenchymal interface or leading edge. These lesions correlated with a select subset of patients, consisting predominantly of young men. CONCLUSIONS Our review identified a distinct pattern of pneumothorax-associated fibroblastic lesions in a subset of cases of spontaneous pneumothorax. Whether this is related to the pathogenesis of the pneumothorax remains to be elucidated.

Alterations in the RB, p16, and cyclin D1 cell cycle control pathway in osteosarcomas

The retinoblastoma (RB) tumor suppressor is believed to play an important role in the pathogenesis of osteosarcomas. Loss of function of RB may occur through abrogation of the RB/p16/cyclin Dl regularity pathway. Our study evaluated the protein expression of pRB, p16, and cyclin D1 in osteosarcomas: 27 tumor specimens from 20 patients were immunostained with antibodies to pRB, p16, and cyclin Dl using the avidin-biotin method with antigen retrieval. In a subset of cases, the immuno-histochemical expression of pRB was correlated with loss of heterozygosity (LOH) at the RB locus; 56% of the specimens (15/27 cases) had an altered component of the pRB Ip16/cyclin D1 pathway with pRB the most frequent abnormal protein (9/27 or 33%) and p16 and cyclin D1 immunohistochemically abnormal in 2 and 3 patients, respectively. Only one protein in the pathway was abnormally expressed in any individual specimen. Absent pRB expression was associated with LOH in 7 of 15 cases (47%), white in another 8 cases pRB was expressed immunohistochemically despite LOH noted by molecular analysis. Two cases were heterozygous by molecular analysis and demonstrated positive nuclear staining. These results support the role of the RB cell cycle control pathway (pRB, p16, cyclin D1) in the pathogenesis of a subset of osteosarcomas. The results also seem to identify a subset of osteosarcomas (about one-third in our study) in which other genetic factors, including others in this pathway, may be important.

Clinical mutational profiling of 1006 lung cancers by next generation sequencing

Analysis of lung adenocarcinomas for actionable mutations has become standard of care. Here, we report our experience using next generation sequencing (NGS) to examine AKT1, BRAF, EGFR, ERBB2, KRAS, NRAS, and PIK3CA genes in 1006 non-small cell lung cancers in a clinical diagnostic setting. NGS demonstrated high sensitivity. Among 760 mutations detected, the variant allele frequency (VAF) was 2–5% in 33 (4.3%) mutations and 2–10% in 101 (13%) mutations. A single bioinformatics pipeline using Torrent Variant Caller, however, missed a variety of EGFR mutations. Mutations were detected in KRAS (36% of tumors), EGFR (19%) including 8 (0.8%) within the extracellular domain (4 at codons 108 and 4 at codon 289), BRAF (6.3%), and PIK3CA (3.7%). With a broader reportable range, exon 19 deletion and p.L858R accounted for only 36% and 26% of EGFR mutations and p.V600E accounted for only 24% of BRAF mutations. NGS provided accurate sequencing of complex mutations seen in 19% of EGFR exon 19 deletion mutations. Doublet (compound) EGFR mutations were observed in 29 (16%) of 187 EGFR-mutated tumors, including 69% with two non-p.L858R missense mutations and 24% with p.L858 and non-p.L858R missense mutations. Concordant VAFs suggests doublet EGFR mutations were present in a dominant clone and cooperated in oncogenesis. Mutants with predicted impaired kinase, observed in 25% of BRAF-mutated tumors, were associated with a higher incidence of concomitant activating KRAS mutations. NGS demonstrates high analytic sensitivity, broad reportable range, quantitative VAF measurement, single molecule sequencing to resolve complex deletion mutations, and simultaneous detection of concomitant mutations.Analysis of lung adenocarcinomas for actionable mutations has become standard of care. Here, we report our experience using next generation sequencing (NGS) to examine AKT1, BRAF, EGFR, ERBB2, KRAS, NRAS, and PIK3CA genes in 1006 non-small cell lung cancers in a clinical diagnostic setting. NGS demonstrated high sensitivity. Among 760 mutations detected, the variant allele frequency (VAF) was 2-5% in 33 (4.3%) mutations and 2-10% in 101 (13%) mutations. A single bioinformatics pipeline using Torrent Variant Caller, however, missed a variety of EGFR mutations. Mutations were detected in KRAS (36% of tumors), EGFR (19%) including 8 (0.8%) within the extracellular domain (4 at codons 108 and 4 at codon 289), BRAF (6.3%), and PIK3CA (3.7%). With a broader reportable range, exon 19 deletion and p.L858R accounted for only 36% and 26% of EGFR mutations and p.V600E accounted for only 24% of BRAF mutations. NGS provided accurate sequencing of complex mutations seen in 19% of EGFR exon 19 deletion mutations. Doublet (compound) EGFR mutations were observed in 29 (16%) of 187 EGFR-mutated tumors, including 69% with two non-p.L858R missense mutations and 24% with p.L858 and non-p.L858R missense mutations. Concordant VAFs suggests doublet EGFR mutations were present in a dominant clone and cooperated in oncogenesis. Mutants with predicted impaired kinase, observed in 25% of BRAF-mutated tumors, were associated with a higher incidence of concomitant activating KRAS mutations. NGS demonstrates high analytic sensitivity, broad reportable range, quantitative VAF measurement, single molecule sequencing to resolve complex deletion mutations, and simultaneous detection of concomitant mutations.

Pleural Fluid Cytology of the Polymorphous Variant of EBV-Positive Diffuse Large B-Cell Lymphoma: First Report and Distinction from a Reactive Process

EBV-positive diffuse large B-cell lymphoma of the elderly is a newly described aggressive lymphoma predominantly affecting patients >50 years of age. Patients may present with nodal and/or extranodal involvement. The lung is one of the more common extranodal sites. The incidence of pleural fluid involvement is less well described. In one study by Oyama et al., pleural effusions were noted in nine percent of cases. Identification of pleural fluid involvement could be important as it may carry prognostic importance in staging (it typically occurs more often in cases with widespread disease), and it could be a relatively easy means of establishing a diagnosis in newly presenting cases. We report the first description of the pleural fluid cytology in a case of EBV-positive diffuse large B-cell lymphoma of the elderly.