Luca Cima

Validation of 34betaE12 immunoexpression in clear cell papillary renal cell carcinoma as a sensitive biomarker

Clear cell papillary renal cell carcinoma (CCPRCC) is a recently recognised neoplasm with a broad spectrum of morphological characteristics, thus representing a challenging differential diagnosis, especially with the low malignant potential multicystic renal cell neoplasms and clear cell renal cell carcinoma. We selected 14 cases of CCPRCC with a wide spectrum of morphological features diagnosed on morphology and CK7 immunoreactivity and analysed them using a panel of immunohistochemical markers, focusing on 34βE12 and related CKs 1,5,10 and 14 and several molecular analyses such as fluorescence in situ hybridisation (FISH), array comparative genomic hybridisation (aCGH), VHL methylation, VHL and TCEB1 sequencing and multiplex ligation-dependent probe amplification (MLPA). Twelve of 13 (92%) CCPRCC tumours were positive for 34βE12. One tumour without 3p alteration by FISH revealed VHL mutation and 3p deletion at aCGH; thus, it was re-classified as clear cell RCC. We concluded that: (1) immunohistochemical expression of CK7 is necessary for diagnostic purposes, but may not be sufficient to identify CCPRCC, while 34βE12, in part due to the presence of CK14 antigen expression, can be extremely useful for the recognition of this tumour; and (2) further molecular analysis of chromosome 3p should be considered to support of CCPRCC diagnosis, when FISH analysis does not evidence the common loss of chromosome 3p.

Reprofiling Metastatic Samples for Chromosome 9p and 14q Aberrations as a Strategy to Overcome Tumor Heterogeneity in Clear-cell Renal Cell Carcinoma.

Losses of chromosomes 9p and 14q are associated with worse outcomes in patients affected by clear-cell renal cell carcinoma (RCC) and are helpful for prognostic risk stratification. Both chromosomal loci harbor several hot-spot molecular pathways suitable for targeted therapeutic interventions. Intratumor heterogeneity may foster tumor adaptation and therapeutic failure. We sought to investigate the presence of losses of the hot spots of chromosomal loci 9p and 14q in primary clear-cell RCC and matched metastatic tissues. CD10 and CD13 were performed on 7 cases of clear-cell RCC with hematogenous tissue metastases. Cytogenetic fluorescence in situ hybridization analysis was performed on primary and matched metastatic tissues using specific probes mapping the 9p and the 14q loci. The loss of chromosome 9p was observed in 85% of both primary clear-cell RCCs and in matched metastases; 14% showed discordance between primary and matched metastases showing gains. The loss of chromosome 14q was observed in 58% of both primary and matched metastases. Only 3/7 (42%) did show an equal status of loss of chromosome 14q. Heterogeneity of the cytogenetic status between metastatic and primary clear-cell RCCs is observed for the loss of chromosome 14q rather than chromosome 9p. The impact of chromosome 14q cytogenetic status, harboring the HIF1 gene, a major driver for the angiogenenic switch, may drive the efficacy of targeted inhibitors, whereas the loss of chromosome 9p, harboring other hot-spot genes, seems to be related to the metastatic behavior per se, without cytogenetic modulation. Reprofiling the metastatic tissue, as compared with the primary tumor, in patients affected by metastatic RCC could be a novel approach to overcome resistance to VEGF(Rs)-targeting agents.

Pediatric optic nerve sheath meningioma.

W e read with great interest the report by Nabavizadeh et al (1) regarding the rare occurrence and aggressive behavior of optic nerve sheath meningioma in the pediatric population (2,3). We had the opportunity to evaluate a 10-year-old girl with a 2-month history of progressive left proptosis. Brain magnetic resonance image (MRI) demonstrated a left intraorbital mass with thickening of the orbital portion of the optic nerve with intense and heterogeneous contrast enhancement. The provisional diagnosis was optic nerve glioma. Two months later, MRI showed enlargement of the mass with extension to the orbital apex but without evidence of intracranial invasion (Fig. 1). Because of rapid tumor growth, the patient underwent frontotemporal craniotomy for tumor excision. Histopathology of the specimen showed fragments of optic nerve surrounded and infiltrated by round aggregates of cells with eosinophilic cytoplasm, round-to-oval uniform nuclei with some pseudoinclusions. No necrosis and mitotic figures were found. Immunohistochemical staining revealed diffuse cytoplasmic epithelial membrane antigen expression, confirming a meningothelial origin (Fig. 2). Progesterone receptor, glial fibrillary acidic protein, leukocyte common antigen, and S-100 were negative. Ki-67 labeling index was 6%–7%, and Bcl-2 labeling index was 15%–20%. The final diagnosis was meningothelial meningioma, WHO Grade I, with recommendation of careful follow-up because of the Ki-67 index. No recurrence of disease was documented on MRI 3 months later. Clinical behavior of primary optic nerve sheath meningioma seems more aggressive in pediatric patients than in adults, with rapid visual decline and greater likelihood of intracranial extension and recurrence. Although established

Primary cutaneous B-cell lymphoma and chronic leg ulcers in a patient with type 2 diabetes

The incidences of type 2 diabetes mellitus and many cancers are rapidly increasing worldwide. Diabetes is a strong risk factor for some cancers (including lymphomas) and is also associated with adverse cancer outcomes. After gastrointestinal tract, the skin is the second most frequent extranodal site involved by non-Hodgkin lymphomas and the cutaneous B-cell lymphomas (CBCLs) range from 25% to 30% of all primary cutaneous lymphomas. The primary cutaneous diffuse large B-cell lymphoma (PCDLBCL) is an aggressive lymphoma with a poor prognosis, representing roughly 20% of all primary CBCLs. Classically, the cutaneous manifestation of this lymphoma is a red or violaceous tumors arising on a leg. To date, despite the large body of evidence suggesting that diabetes is strongly associated with an increased risk of some cancers, very little information is available regarding a possible association between type 2 diabetes and primary cutaneous diffuse large B-cell lymphoma. In this report, we will present the case of a white adult patient with type 2 diabetes with chronic leg ulcers complicated by a primary cutaneous diffuse large B-cell lymphoma. Learning points: Diabetes mellitus is increasing worldwide as well as the incidence of many cancers. Diabetes mellitus is a powerful risk factor for some cancers (including lymphomas) and is strongly associated with adverse cancer outcomes. Seen that diabetes is strongly associated with an increased risk of cancers (including cutaneous lymphomas), clinicians should always keep in mind this complication in elderly patients with type 2 diabetes, even in a chronic leg ulcer with hypertrophy of the wound edge, which is hard to heal and does not have the typical characteristics of a diabetic or vascular ulcer. In these cases, a biopsy should be performed to rule out a neoplasm. Early diagnosis and correct management of cancer in a patient with type 2 diabetes are crucial to improve clinical outcomes.

Next-generation repeat-free FISH probes for DNA amplification in glioblastoma in vivo: Improving patient selection to MDM2-targeted inhibitors.

A next-generation FISH probe mapping to the MDM2 locus-specific region has recently been designed. The level of MDM2 gene amplification (high versus low) may allow selection of patients for cancer treatment with MDM2 inhibitors and may predict their responsiveness. We investigated the spectrum of MDM2 gene alterations using the new probes in vivo after visualizing single neoplastic cells in situ from a series of glioblastomas. Signals from next-generation repeat-free FISH interphase probes were identified in tissue microarrays that included 3 spots for each of the 48 cases. The murine double minutes (MDM2)-specific DNA probe and the satellite enumeration probe for chromosome 12 were used. Three cases (6%) showed more than 25 signals (high gene amplification), and 7 (15%) showed 3-10 signals (gains); among these, 4 cases (8%) had an equal number of MDM2 and centromeric signals on chromosome 12 (polyploidy). Genomic heterogeneity was observed only in 3 cases with low gene amplification. In our series, 6% of glioblastomas exhibited high MDM2 amplification (in vivo) with a pattern related to the known double minutes/chromothripsis phenomenon (in situ), and only cases with low amplification showed genomic heterogeneity. We concluded that the rate of MDM2 gene amplification can be a useful predictive biomarker to improve patient selection.

Liver Metastases From Renal Oncocytoma With Vascular Extension

The 2016 World Health Organization Renal Tumor Classification defines renal oncocytoma (RO) as a benign epithelial tumor; however, malignant histopathologic features have been documented. Rare cases with metastases have been reported. We describe the case of a 62-year-old woman who was referred to the Urology Clinic for a routine work-up. Magnetic resonance imaging and computerized tomography showed a 7-cm mass in the middle and lower portions of the left kidney and 2 suspected liver metastases. The patient underwent surgery. Microscopically both renal and liver lesions presented solid, solid-nested, and microcystic architecture, composed predominantly of large eosinophilic cells without any worrisome pattern except the vascular extension. The cells were positive for S100A1, CD117, and PAX-8 and negative for CAIX, CK7, and AMACR. Fluorescence in situ hybridization showed a disomic profile for the chromosomes 1, 2, 6, 7, 10, 17. No mutation of coding sequence of the SDHB, SDHC, SDHD, VHL, and BHD genes and no loss of heterozygosity at 3p were found. The final diagnosis was “RO” according to the 2016 World Health Organization Renal Tumor Classification with “liver metastases.” This report provides a wide clinical-pathologic, immunophenotypical and molecular documentation of a RO with liver metastases.

Oil-Red-O is a useful tool to assess donor liver steatosis on frozen sections during transplantation

Oil Red O is a useful tool to assess donor liver steatosis on frozen sections during transplantation. Steatosis is a frequent finding in liver evaluation during transplantation, accounting for 9% to 26% of biopsied donor liver. The degree of macrovesicular steatosis is classified as mild, moderate, and severe; the latter is considered an absolute contraindication to liver transplantation because it is associated with poor allograft outcome. Because of the scarcity of organs, there is a debate whether livers with less severe macrovesicular steatosis are still suitable for transplant. Consequently, tools or methods that allow a more accurate intraoperative assessment of steatosis on frozen sections are mandatory. The aim of this study is to improve intraoperative evaluation of steatosis during transplantation using Oil Red O stain on liver biopsies. METHODS Twenty consecutive liver biopsies of donors were collected during transplantation procedures from September 2017 to February 2018 at the Institute of Pathology of the University and Hospital Trust of Verona, Italy. Each liver biopsy was cut at a different thickness (3, 5, and 8 μm) and stained with both Oil Red O and conventional hematoxylin and eosin for intraoperative consultation. The degree (percentage of hepatocytes involved) of fatty changes was recorded. The results obtained during the intraoperative consultation were finally compared with the formalin-fixed and paraffin-embedded permanent section. RESULTS Assessment of steatosis on hematoxylin and eosin frozen sections was reported as mild in 17 cases (85%), moderate in 2 cases (10%) and severe in 1 case (5%). Oil Red O frozen sections reported the following results: mild steatosis in 16 cases (80%), moderate in 2 cases (10%), and severe in 2 cases (10%). The percentage of liver steatosis obtained with Oil Red O was consistent in all cases with that of the permanent sections. The staining procedure for Oil Red O required approximately 18 minutes. CONCLUSIONS Oil Red O special stain is a fast and inexpensive tool to improve the assessment of steatosis on frozen biopsies during liver transplantation.

Low-grade neuroepithelial tumor: Unusual presentation in an adult without history of seizures: LGNT in an adult

Low‐grade neuroepithelial tumors (LGNT) show a broad histopathological spectrum and may be difficult to classify using current World Health Organization (WHO) criteria. A 57‐year‐old man came to medical attention because of headaches. The patient medical history was otherwise unremarkable. Magnetic resonance imaging (MRI) revealed a 2.5 cm lesion, partially cystic, with an increased signal on T2‐weighted imaging, located in the right frontal lobe. The patient underwent right frontal craniotomy and the surgical specimen was entirely evaluated. Microscopic examination showed a tumor arranged predominantly in sheets and nests, with an infiltrative growth pattern and oligodendroglioma‐like appearance. Tumor cells were round to oval with cytoplasmic clearing, hyperchromatic nuclei and inconspicuous nucleoli. Only one mitosis was identified. Necrosis was absent. Differential diagnostic considerations included oligodendroglioma, clear cell ependymoma, polymorphous low‐grade neuroepithelial tumor of the young (PLNTY) and long‐term epilepsy‐associated tumor with clear cell morphology. Neoplastic cells showed positivity for glial fibrillary acidic protein (GFAP), oligodendrocyte transcription factor 2 (OLIG2), α‐thalasemia X‐linked mental retardation syndrome (ATRX) (retained nuclear expression) and CD34. Epithelial membrane antigen (EMA), neuronal nuclear antigen, microtubule‐associated protein‐2e, cyclo‐oxygenase‐2, chromogranin A and isocitrate dehydrogenase 1 (IDH1) (R132H) were negative. Ki‐67 labeling index was 2–3%. Molecular analysis identified neither IDH1/IDH2 mutations nor 1p19q codeletion. Rapidly accelerated fibrosarcoma homolog B1 (BRAF) V600E mutation was also absent by both molecular and immunohistochemical testing. Polymerase chain reaction analysis revealed the presence of fibroblast growth factor receptor 3 (FGFR3)‐transforming acidic coiled‐coil (TACC) fusion. Taken together, the morphological, immunohistochemical and molecular findings supported the final diagnosis of PLNTY.

Validation of remote digital frozen sections for cancer and transplant intraoperative services

Introduction: Whole-slide imaging (WSI) technology can be used for primary diagnosis and consultation, including intraoperative (IO) frozen section (FS). We aimed to implement and validate a digital system for the FS evaluation of cancer and transplant specimens following recommendations of the College of American Pathologists. Materials and Methods: FS cases were routinely scanned at ×20 employing the “Navigo” scanner system. IO diagnoses using glass versus digital slides after a 3-week washout period were recorded. Intraobserver concordance was evaluated using accuracy rate and kappa statistics. Feasibility of WSI diagnoses was assessed by the way of sensitivity, specificity, as well as positive and negative predictive values. Participants also completed a survey denoting scan time, time spent viewing cases, preference for glass versus WSI, image quality, interface experience, and any problems encountered. Results: Of the 125 cases submitted, 121 (436 slides) were successfully scanned including 93 oncological and 28 donor-organ FS biopsies. Four cases were excluded because of failed digitalization due to scanning problems or sample preparation artifacts. Full agreement between glass and digital-slide diagnosis was obtained in 90 of 93 (97%, κ = 0.96) oncology and in 24 of 28 (86%, κ = 0.91) transplant cases. There were two major and one minor discrepancy for cancer cases (sensitivity 100%, specificity 96%) and two major and two minor disagreements for transplant cases (sensitivity 96%, specificity 75%). Average scan and viewing/reporting time were 12 and 3 min for cancer cases, compared to 18 and 5 min for transplant cases. A high diagnostic comfort level among pathologists emerged from the survey. Conclusions: These data demonstrate that the “Navigo” digital WSI system can reliably support an IO FS service involving complicated cancer and transplant cases.

Virtual autopsy as a screening test before traditional autopsy: The verona experience on 25 Cases

Background: Interest has grown into the use of multidetector computed tomography (CT) and magnetic resonance imaging as an adjunct or alternative to the invasive autopsy. We sought to investigate these possibilities in postmortem CT scan using an innovative virtual autopsy approach. Methods: Twenty-five postmortem cases were scanned with the Philips Brilliance CT-64 and then underwent traditional autopsy. The images were interpreted by two blinded forensic pathologists assisted by a radiologist with the INFOPSY® Digital Autopsy Software System which provides three-dimensional images in Digital Imaging and Communications in Medicine format. Diagnostic validity of virtual autopsy (accuracy rate, sensitivity, specificity, and predictive values) and concordance between the two forensic pathologists (kappa intraobserver coefficients) were determined. Results: The causes of death at traditional autopsies were hemorrhage due to traumatic injuries (n = 8), respiratory failure (5), asphyxia due to drowning (4), asphyxia due to hanging or strangulation (2), heart failure (2), nontraumatic hemorrhage (1), and severe burns (1). In two cases, the cause of death could not be ascertained. In 15/23 (65%) cases, the cause of death diagnosed after virtual autopsy matched the diagnosis reported after traditional autopsy. In 8/23 cases (35%), traditional autopsy was necessary to establish the cause of death. Digital data provided relevant information for inferring both cause and manner of death in nine traumatic cases. The validity of virtual autopsy as a diagnostic tool was higher for traumatic deaths than other causes of death (accuracy 84%, sensitivity 82%, and specificity 86%). The concordance between the two forensic pathologists was almost perfect (>0.80). Conclusions: Our experience supports the use of virtual autopsy in postmortem investigations as an alternative diagnostic practice and does suggest a potential role as a screening test among traumatic deaths.