Ema Dragoescu

Preservation of Fine-Needle Aspiration Specimens for Future Use in RNA-Based Molecular Testing

The application of ancillary molecular testing is becoming more important for the diagnosis and classification of disease. The use of fine‐needle aspiration (FNA) biopsy as the means of sampling tumors in conjunction with molecular testing could be a powerful combination. FNA is minimally invasive, cost effective, and usually demonstrates accuracy comparable to diagnoses based on excisional biopsies. Quality control (QC) and test validation requirements for development of molecular tests impose a need for access to pre‐existing clinical samples. Tissue banking of excisional biopsy specimens is frequently performed at large research institutions, but few have developed protocols for preservation of cytologic specimens. This study aimed to evaluate cryopreservation of FNA specimens as a method of maintaining cellular morphology and ribonucleic acid (RNA) integrity in banked tissues.

Dual Action of miR-125b As a Tumor Suppressor and OncomiR-22 Promotes Prostate Cancer Tumorigenesis

MicroRNAs (miRs) are a novel class of small RNA molecules, the dysregulation of which can contribute to cancer. A combinatorial approach was used to identify miRs that promote prostate cancer progression in a unique set of prostate cancer cell lines, which originate from the parental p69 cell line and extend to a highly tumorigenic/metastatic M12 subline. Together, these cell lines are thought to mimic prostate cancer progression in vivo. Previous network analysis and miR arrays suggested that the loss of hsa-miR-125b together with the overexpression of hsa-miR-22 could contribute to prostate tumorigenesis. The dysregulation of these two miRs was confirmed in human prostate tumor samples as compared to adjacent benign glandular epithelium collected through laser capture microdissection from radical prostatectomies. In fact, alterations in hsa-miR-125b expression appeared to be an early event in tumorigenesis. Reverse phase microarray proteomic analysis revealed ErbB2/3 and downstream members of the PI3K/AKT and MAPK/ERK pathways as well as PTEN to be protein targets differentially expressed in the M12 tumor cell compared to its parental p69 cell. Relevant luciferase+3’-UTR expression studies confirmed a direct interaction between hsa-miR-125b and ErbB2 and between hsa-miR-22 and PTEN. Restoration of hsa-miR-125b or inhibition of hsa-miR-22 expression via an antagomiR resulted in an alteration of M12 tumor cell behavior in vitro. Thus, the dual action of hsa-miR-125b as a tumor suppressor and hsa-miR-22 as an oncomiR contributed to prostate tumorigenesis by modulations in PI3K/AKT and MAPK/ERK signaling pathways, key pathways known to influence prostate cancer progression.

miR-9 Acts as an OncomiR in Prostate Cancer through Multiple Pathways That Drive Tumour Progression and Metastasis

Identification of dysregulated microRNAs (miRNAs) in prostate cancer is critical not only for diagnosis, but also differentiation between the aggressive and indolent forms of the disease. miR-9 was identified as an oncomiR through both miRNA panel RT-qPCR as well as high-throughput sequencing analysis of the human P69 prostate cell line as compared to its highly tumorigenic and metastatic subline M12, and found to be consistently upregulated in other prostate cell lines including DU-145 and PC3. While miR-9 has been characterized as dysregulated either as an oncomiR or tumour suppressor in a variety of other cancers including breast, ovarian, and nasopharyngeal carcinomas, it has not been previously evaluated and proven as an oncomiR in prostate cancer. miR-9 was confirmed an oncomiR when found to be overexpressed in tumour tissue as compared to adjacent benign glandular epithelium through laser-capture microdissection of radical prostatectomy biopsies. Inhibition of miR-9 resulted in reduced migratory and invasive potential of the M12 cell line, and reduced tumour growth and metastases in male athymic nude mice. Analysis showed that miR-9 targets e-cadherin and suppressor of cytokine signalling 5 (SOCS5), but not NF-ĸB mRNA. Expression of these proteins was shown to be affected by modulation in expression of miR-9.

Small cell osteosarcoma with Ewing sarcoma breakpoint region 1 gene rearrangement detected by interphase fluorescence in situ hybridization.

Because of its characteristic morphologic appearance, small cell osteosarcoma (SCO) can be confused with other small round cell malignancies of the bone, most importantly with Ewing sarcoma, making this distinction difficult. A specific tool used in separating SCO from Ewing sarcoma has been the detection of Ewing sarcoma breakpoint region 1 (EWSR1) gene rearrangements in Ewing sarcoma and their absence in SCO. However, there are rare case reports that have documented the existence of EWSR1 gene rearrangement in SCO. In this report, we describe another case of SCO with an EWSR1 gene rearrangement detected by interphase fluorescence in situ hybridization. Our finding adds support to the existing evidence that SCO is a tumor that can be characterized by EWSR1 gene arrangements. Therefore, we caution the pathology community not to rely solely on molecular studies in distinguishing SCO from Ewing sarcoma.

Renal cell carcinoma, unclassified with medullary phenotype: poorly differentiated adenocarcinomas overlapping with renal medullary carcinoma

Renal medullary carcinoma (RMC) is a highly aggressive renal cell carcinoma arising in the collecting system and requiring careful correlation with status of sickle cell trait. A panel of international experts has recently proposed provisional diagnostic terminology, renal cell carcinoma, unclassified, with medullary phenotype, based on encountering an extraordinarily rare tumor with RMC morphology and immunophenotype in an individual proven not to have a hemoglobinopathy. Herein, we extend this observation to a cohort of 5 such tumors, morphologically similar to RMC, lacking SMARCB1 expression by immunohistochemistry, but each without evidence of a hemoglobinopathy. The tumors arose in 4 men and 1 woman with a mean age of 44 years, occurring in 3 left and 2 right kidneys. Clinically, aggression was apparent with involvement of perinephric adipose tissue in all 5 cases, nodal metastasis in 4 of 5 cases, and death of disease in 4 of 5 cases within 3-27 months. Histologic sections showed poorly differentiated adenocarcinoma, often with solid and nested growth patterns, as well as infiltrative glandular, tubulopapillary, cribriform, or reticular growth. Rhabdoid and sarcomatoid cytomorphology was seen in a subset. All tumors showed PAX8 nuclear positivity and SMARCB1 loss, with OCT3/4 expression in 4 of 5 cases. In summary, this first series of renal cell carcinoma, unclassified, with medullary phenotype documents tumors with morphologic, immunophenotypic, and prognostic features of RMC occurring in individuals without sickle cell trait. Although greater biologic and molecular understanding is needed, the available evidence points to these cases representing a sporadic counterpart to sickle cell trait-associated RMC.

Indications for renal fine needle aspiration biopsy in the era of modern imaging modalities

Background: Renal fine needle aspiration biopsy (FNAB) has become an uncommon procedure in the era of renal helical computed tomography (CT), which has high diagnostic accuracy in the characterization of renal cortical lesions. This study investigates the current indications for renal FNAB. Having knowledge of the specific clinico-radiologic scenario that led to the FNAB, cytopathologists are better equipped to expand or narrow down their differential diagnosis. Materials and Methods: All renal FNABs performed during a 6 year interval were retrieved. Indication for the procedure was determined from the clinical notes and radiology reports. Results: Forty six renal FNABs were retrieved from 43 patients (14 females and 29 males with a mean age of 52 years [range, 4-81 years]). Twenty one cases (45.6%) were performed under CT-guidance and 25 cases (54.4%) under US-guidance. There were four distinct indications for renal FNAB: (1) solid renal masses with atypical radiological features or poorly characterized on imaging studies due to lack of intravenous contrast or body habitus (30.2%); (2) confirmation of radiologically suspected renal cell carcinoma in inoperable patients (advanced stage disease or poor surgical candidate status) (27.9%); (3) kidney mass in a patient with a prior history of other malignancy (27.9%); and (4) miscellaneous (drainage of abscess, indeterminate cystic lesion, urothelial carcinoma) (14.0%). 36 patients (83.7%) received a specific diagnosis based on renal FNAB cytology. Conclusions: Currently, renal fine needle aspiration remains a useful diagnostic tool in selected clinico-radiologic scenarios.

Acute Appendicitis and Pneumatosis in a Duplicated Appendix With Schistosoma Remnants.

Appendiceal pneumatosis is rare, reported either in the context of acute appendicitis or enterocolitis. Here, we report the case of an elderly adult in whom the acute appendicitis was associated with pneumatosis and occurred in the context of a malformed appendix with pathogenic organism remnants. A 72-year-old man presented with abdominal pain 3 weeks after posttraumatic dorsolumbar surgery. The computed tomography scan showed acute appendicitis and 2 diverticula. On microscopy, the appendix showed acute appendicitis along with a Cave-Wallbridge type A duplication. In addition, several optically clear spaces were observed in the entire appendiceal wall consistent with pneumatosis of the appendix. Focally, calcified structures suggesting pathogenic organisms such as Schistosoma were noted as well. In conclusion, we report a case of appendiceal pneumatosis occurring in the context of acute appendicitis in a duplicated appendix, with presence of calcified structures suggestive of pathogenic organisms.

Cytokeratin 5/6 and P63 immunophenotype of thyroid lymphoepithelial complexes

Thyroid lymphoepithelial complexes (LECos) are rare, being reported in lymphoma, Graves-Basedow disease, Hashimoto thyroiditis, pericarcinomatous thyroid or in the context of branchial cleft-like cysts. Here we report immunohistochemical expression of cytokeratin 5/6, P63 and TTF1 in 6 cases of thyroid LECos. Two cases had carbimazole treatment for hyperthyroidia and Graves disease. Anti-thyroglobulin, -thyroperoxidase or -TSH antibodies were detected in 4 cases. NSAID or poviodone iodine allergy were present in 2 cases. The treatment consisted in total thyroidectomy or lobectomy. Microscopy showed nodular goiter and focal lymphocytic thyroiditis. Basaloid LECos were seen in all thyroids while squamoid LECos in 2. Associated lesions were papillary thyroid microcarcinoma (2 cases), solid cell nest, thyroglosal duct remnant, lymphoepithelial cyst and thymus-parathyroid unit (one case each). Cytokeratin 5/6 was expressed in both squamoid and basaloid LECos along with P63. TTF1 expression was faint or absent. In conclusion LECos may occur in the context of autoimmune thyroiditis or of a specific immune susceptibility background. The expression of CK5/6 and of P63 suggests a squamous differentiation including in the basaloid LECos. The etiologic relevance of these immunostainings remains limited although rather suggestive of a metaplastic process than of migration-abnormalities.

Do Osteoblastoma and Osteoid Osteoma Represent the Same Pathologic Entity

Case:We present the case of a fourteen-year-old boy with a benign diaphyseal femoral lesion that initially fulfilled the diagnostic criteria for osteoid osteoma but rapidly progressed and was finally diagnosed as osteoblastoma. Conclusion:Clinical and radiographic criteria provide imperfect and somewhat arbitrary distinguishing features between osteoid osteoma and osteoblastoma. Although previous work highlighted the importance of histologic interpretation in this regard, recent comparative histologic and immunohistochemical evaluations failed to differentiate the two. We conclude that consideration should be given to the classification of osteoid osteoma and osteoblastoma as the same pathologic entity, with treatment based on presentation along a continuum of aggressiveness, and that future directions for study may include identification of factors predictive of progression and/or recurrence.

Fine Needle Aspiration Cytology of Endocrine Glands

Although fine needle aspiration is considered a relatively recent diagnostic procedure, one of the earliest known reports of this technique was documented in 1847 by M. Kun, who noted that “in this manner a microscopic examination of the tumour can be practised on the living subject, and its nature ascertained before having recourse to an operation” [1]. Determining the nature of an unknown lesion without relying on surgical intervention is the primary indication for fine needle aspiration biopsy (FNAB), which has finally gained widespread acceptance as a diagnostic technique, after many years of lukewarm interest. Drs. Martin and Stewart briefly popularized the technique in the early 1930s at the Memorial Center for Cancer and Allied Diseases in New York [2,3], but it was not until the 1970s that FNAB was practiced with any frequency in the United States. This surge in popularity followed a wave of interest in the technique that began with a group of clinicians in Scandinavia and Europe, and that was accompanied by several improvements in the technique and seminal publications regarding the method [4,5]. By the early 1980s, FNAB had become reasonably well established in the United States [6,7]. Since that time, the technique has seen dramatic increases in use by clinicians, radiologists, and pathologists.