Mariana M. Cajaiba

DICER1 mutations in childhood cystic nephroma and its relationship to DICER1-renal sarcoma.

The pathogenesis of cystic nephroma of the kidney has interested pathologists for over 50 years. Emerging from its initial designation as a type of unilateral multilocular cyst, cystic nephroma has been considered as either a developmental abnormality or a neoplasm or both. Many have viewed cystic nephroma as the benign end of the pathologic spectrum with cystic partially differentiated nephroblastoma and Wilms tumor, whereas others have considered it a mixed epithelial and stromal tumor. We hypothesize that cystic nephroma, like the pleuropulmonary blastoma in the lung, represents a spectrum of abnormal renal organogenesis with risk for malignant transformation. Here we studied DICER1 mutations in a cohort of 20 cystic nephromas and 6 cystic partially differentiated nephroblastomas, selected independently of a familial association with pleuropulmonary blastoma and describe four cases of sarcoma arising in cystic nephroma, which have a similarity to the solid areas of type II or III pleuropulmonary blastoma. The genetic analyses presented here confirm that DICER1 mutations are the major genetic event in the development of cystic nephroma. Further, cystic nephroma and pleuropulmonary blastoma have similar DICER1 loss of function and ‘hotspot’ missense mutation rates, which involve specific amino acids in the RNase IIIb domain. We propose an alternative pathway with the genetic pathogenesis of cystic nephroma and DICER1-renal sarcoma paralleling that of type I to type II/III malignant progression of pleuropulmonary blastoma.

GLUT1 expression in malignant tumors and its use as an immunodiagnostic marker

OBJECTIVE: To analyze glucose transporter 1 expression patterns in malignant tumors of various cell types and evaluate their diagnostic value by immunohistochemistry. INTRODUCTION: Glucose is the major source of energy for cells, and glucose transporter 1 is the most common glucose transporter in humans. Glucose transporter 1 is aberrantly expressed in several tumor types. Studies have implicated glucose transporter 1 expression as a prognostic and diagnostic marker in tumors, primarily in conjunction with positron emission tomography scan data. METHODS: Immunohistochemistry for glucose transporter 1 was performed in tissue microarray slides, comprising 1955 samples of malignant neoplasm from different cell types. RESULTS: Sarcomas, lymphomas, melanomas and hepatoblastomas did not express glucose transporter 1. Forty-seven per cent of prostate adenocarcinomas were positive, as were 29% of thyroid, 10% of gastric and 5% of breast adenocarcinomas. Thirty-six per cent of squamous cell carcinomas of the head and neck were positive, as were 42% of uterine cervix squamous cell carcinomas. Glioblastomas and retinoblastomas showed membranous glucose transporter 1 staining in 18.6% and 9.4% of all cases, respectively. Squamous cell carcinomas displayed membranous expression, whereas adenocarcinomas showed cytoplasmic glucose transporter 1 expression. CONCLUSION: Glucose transporter 1 showed variable expression in various tumor types. Its absence in sarcomas, melanomas, hepatoblastomas and lymphomas suggests that other glucose transporters mediate the glycolytic pathway in these tumors. The data suggest that glucose transporter 1 is a valuable immunohistochemical marker that can be used to identify patients for evaluation by positron emission tomography scan. The function of cytoplasmic glucose transporter 1 in adenocarcinomas must be further examined.

MT1G Hypermethylation: A Potential Prognostic Marker for Hepatoblastoma

Hepatoblastoma comprises only 1% of all cancers in childhood. Because of its low frequency, a small number of prognostic factors are described in hepatoblastoma and most of them are related to resectability. Microarray studies showed a large number of underexpressed genes in hepatoblastoma. Because aberrant DNA methylation has been recognized as an alternative mechanism for tumor suppressor gene inactivation, this could be involved with gene downregulation in these tumors. Despite the rarity of hepatoblastoma, this study evaluated the methylation pattern of 25 genes in 20 paraffin-embedded tumor specimens and five non-neoplastic liver samples (normal control) by quantitative methylation-specific PCR (QMSP). The examination of the methylation profile of hepatoblastoma samples and normal liver specimens revealed a high tumor-specific DNA hypermethylation in the promoter regions of five genes (APC, CDH1, MT1G, RASSF1A, and SOCS1). Furthermore, MT1G hypermethylation showed a significant correlation with poor prognosis of patients with hepatoblastoma. This study represents the first quantitative evaluation of promoter hypermethylation in hepatoblastoma and demonstrated that aberrant methylation is a frequent event in this malignancy. Furthermore, our data provide evidence that MT1G hypermethylation may be useful as prognostic indicator for this disease and suggest that patients with hepatoblastoma may benefit from demethylating drug treatments.

Rhabdomyosarcoma, Wilms tumor, and deletion of the patched gene in Gorlin syndrome

Background A 5-year-old year girl with a medical history of mental retardation, physical abnormalities and a known interstitial deletion of chromosome 9q22–q32 presented with a palpable suprapubic mass. During ultrasound investigation, a left renal mass was also detected. The patient underwent surgical removal of both neoplasms, which were diagnosed as a rhabdomyosarcoma and a Wilms tumor. Seven years later, she presented with macroglossia and a benign mandibular cyst.Investigations Physical examination, karyotyping, abdominal and pelvic ultrasound, brain CT scan, anatomic pathology analysis with immunohistochemistry, and typing of polymorphic markers in the patched (PTCH) gene region.Diagnosis Gorlin syndrome with synchronous rhabdomyosarcoma and Wilms tumor.Management Left nephrectomy, excision of paravesical tumor, excision of mandibular cysts, chemotherapy, and radiotherapy.

Bilateral prepubertal testicular biopsies predict significance of cryptorchidism-associated mixed testicular atrophy, and allow assessment of fertility

IntroductionMixed atrophy of the testis (MAT), a frequent finding in biopsies of formerly cryptorchid and/or infertile patients, is defined as the synchronous occurrence of both seminiferous tubules containing germ cells and Sertoli cell only-tubules in variable proportions. In tubules containing germ cells, different types of abnormalities in spermatogenesis may be seen. The presence of adult spermatids in the biopsy, even in small numbers, correlates with successful spermatozoa retrieval for “in vitro” fertilization techniques. Currently, it is unknown whether precursor lesions of MAT can be identified in cryptorchid patients during childhood. Material and MethodsEighteen formerly cryptorchid adults who had undergone testicular biopsies in childhood had a repeat testicular biopsy to evaluate infertility. In prepubertal biopsies, abnormalities of the testicular parenchyma were classified into types I (slight alterations), II (marked germinal hypoplasia), and III (severe germinal hypoplasia). In postpubertal biopsies, the percentage of tubules containing germ cells and Sertoli cell only-tubules were estimated, as well as the presence of complete spermatogenesis. Abnormalities in spermatogenesis were classified into lesions of the adluminal or basal compartments of seminiferous tubules. ResultsComparison between prepubertal and postpubertal biopsies revealed that most specimens developing from type III lesions presented with incomplete spermatogenesis (P<0.0001) and more severe lesions of the germinal epithelium (P=0.049). DiscussionType III lesions correlated with MAT characteristics that confer a worse prognosis for in vitro fertilization. Thus, MAT characteristics may be predicted in prepubertal cryptorchid patients, allowing a fertility prognosis. The pathogenesis of these lesions, and their possible inclusion into the spectrum of the testicular dysgenesis syndrome, are discussed.

Expanding the clinical spectrum of Frasier syndrome

Frasier syndrome is an uncommon genetic disorder featuring progressive glomerulopathy, male pseudohermaphroditism, and gonadal dysgenesis with increased risk of gonadoblastoma and malignant germ cell tumors. It is caused by mutations in the donor splice site in intron 9 of the WT1 gene. However, because of its rarity there is limited literature available on the precise spectrum and recommended treatment modalities of this syndrome. We present the clinicopathological findings in 4 patients: 3 phenotypically female adolescents presenting with proteinuria and primary amenorrhea and a 6-month-old baby girl presenting with nephrotic syndrome in whom this very unusual case of early onset was confirmed by molecular studies. The significance of early recognition of Frasier syndrome and its differentiation from Denys-Drash syndrome is reviewed and discussed. Our observation of a case presenting with early clinical manifestations, in contrast with the classical presentation in adolescence, justifies the expansion of the clinical spectrum of Frasier syndrome and contributes to the understanding and appropriate clinical management of these patients.

ALK-rearranged renal cell carcinomas in children

Knowledge of the clinicopathological and molecular spectrum of pediatric renal cell carcinomas (RCC) remains limited, and approximately 16%–24% of these neoplasms cannot be classified into specific subtypes. In this review of 168 pediatric RCC prospectively registered on Childrens Oncology Group AREN03B2 protocol, six RCC (3.5%) that demonstrated a unique epithelioid morphology and a peculiar immunophenotypic profile that includes expression of ALK, TFE3, and retention of INI1 was identified. Further investigation revealed ALK rearrangements in all cases, manifested molecularly by fusion transcripts of either VCL‐ALK (3 patients all with sickle cell trait which had been previously reported) or TPM3‐ALK (3 patients, none with sickle cell trait). Based on the shared unique morphologic, immunophenotypic, and genetic features, it was proposed that these neoplasms belonged to a distinct subgroup of RCC frequently occurring in pediatric patients, which they have termed as ALK‐rearranged RCC. Importantly, additional therapeutic options may be available for these patients.

Pediatric cystic nephromas: distinctive features and frequent DICER1 mutations

Cystic nephromas (CNs) are uncommon benign renal neoplasms that present with a bimodal age distribution, affecting either infants/young children or adult females. Although differences between these age groups have been suggested, large studies of pediatric CN have not been conducted. As a result, the nomenclature and diagnostic criteria for these lesions remain controversial. In addition, the morphological overlap seen between CN and cystic partially differentiated nephroblastoma (CPDN) can result in diagnostic dilemmas. This study reviews the morphologic and radiographic features of 44 pediatric CN prospectively enrolled on a Childrens Oncology Group protocol from 2007 to 2013. Although the typical multicystic architecture with thin septa described in adult CN was present in all of our pediatric cases, differences were also identified. We report distinctive features that add to the morphological spectrum of CN in children. Of the 44 cases, 16 had been previously analyzed and reported for DICER1 mutation, and either loss of function or missense mutations or both were identified in 15 of 16. In contrast, we analyzed 10 cases of adult CN, and all were negative for DICER1 mutations; similarly, 6 CPDNs previously analyzed and reported were negative for DICER1 mutations. Therefore, the clinical, morphological, and genetic differences between pediatric and adult CN, as well as between CN and CPDN, suggest that these 3 lesions represent distinct entities.

Expanding the spectrum of ALK-rearranged renal cell carcinomas in children: Identification of a novel HOOK1-ALK fusion transcript.

We have recently characterized the morphologic, immunohistochemical, and molecular features of a distinctive group of renal cell carcinomas (RCC) occurring in children and characterized by ALK gene rearrangements (Cajaiba et al., 2016). These tumors manifested either VCL-ALK (three patients) or TPM3-ALK (three patients) fusion transcripts. The histopathological, ultrastructural, and immunohistochemical features seen in these six cases were unique and no differences were noted among cases with the two different ALK fusion partners. All tumors were composed of cells with eosinophilic cytoplasm and high grade nuclear features arranged in a predominantly solid pattern with a prominent vascular background, showed frequent intracytoplasmic vacuoles, were associated with a striking lymphoplasmacytic infiltrate, and showed a similar immunohistochemical profile with expression of epithelial markers, TFE3 and retention of INI-1. Based on these shared features, as well as their overall prevalence within a large series of pediatric RCC, we proposed that these neoplasms belong to a distinct subgroup of RCC frequently occurring in pediatric patients, which we have referred to as ALK-rearranged RCC. Following the publication of our study, we have identified an additional example of ALK-rearranged RCC in a pediatric patient. This patient was enrolled in the AREN03B2 Children’s Oncology Group protocol with a diagnosis of RCC, and a full set of H&E slides and the institutional pathology report were submitted for central pathology review. The patient, a 16 years old Caucasian male, presented with hematuria and a renal mass, and underwent right radical nephrectomy. There was no history of sickle cell disease or trait, which was present in 3/6 of the previously reported cases. Gross examination revealed a 5.5 cm wellcircumscribed, soft and friable tan mass with areas of hemorrhage. Histological sections demonstrated a neoplasm entirely surrounded by a fibrous pseudocapsule and predominantly composed of papillary architecture (Fig. 1A) alternating with solid areas. The cells were round to polygonal with variable amounts of eosinophilic cytoplasm (Figs. 1B and 1C). Frequent intracytoplasmic lumina were noted, and the presence of dense eosinophilic cytoplasm in a subset of cells was reminiscent of intracytoplasmic inclusions (Figs. 1B and 1C). Scattered mucin-filled glandular structures were also appreciated (Figs. 1 B and 1D). Most of the nuclei were irregular with vesicular chromatin and small nucleoli, consistent with an ISUP Grade 3, and a small subset of cells showed greater degrees of nuclear pleomorphism, hyperchromasia, and more prominent nucleoli consistent with an ISUP Grade 4. A patchy mild lymphoplasmacytic inflammatory infiltrate permeated the tumor cells throughout the neoplasm, and occasional dystrophic calcifications were appreciated. Immunohistochemical stains showed diffuse and strong expression of EMA, vimentin, and TFE3 (nuclear), focal expression of cytokeratin 7, retained INI-1 expression, and lack of p53 accumulation. A diagnosis of MiT family translocation RCC was initially considered, however, FISH studies for a TFE3 rearrangement were negative. Despite the extensive papillary architecture, features overlapping with our previously reported ALK-rearranged RCC were noted, prompting us to investigate for the presence of an ALK fusion transcript in this neoplasm. RNA was extracted from archival formalin-fixed paraffin-embedded (FFPE) tissue as previously described, and in-depth molecular analysis using next generation sequencing (NGS) methodology was then performed in this case in an attempt to identify an ALK fusion transcript, using previously described methodology (Cajaiba et al., 2016). The results demonstrated the presence of a HOOK1ALK fusion transcript spanning exon 20 of HOOK1

Hepatoblastomas and Liver Development: A Study of Cytokeratin Immunoexpression in Twenty-Nine Hepatoblastomas:

Hepatoblastomas (HBs) recapitulate liver development. It is possible that HBs result from malignant transformation of hepatic precursor cells, and they may reflect a blockage in normal development. Here we study the expression of cytokeratins (CKs) in order to delineate the immunoprofile and relationship with liver development, as well as vimentin and alphafetoprotein (AFP), of HBs. Immunohistochemistry was performed in a tissue microarray (TMA) containing representative areas of 18 HBs (fetal and/or embryonal and/or mesenchymal); we also reviewed 11 cases not included in the TMA. No cases stained for CKs 1, 5/6, 7, 10, 13, 15, 16, 20, and 34βE12. CK8 stained 73.07% of fetal, 50% of embryonal, and 18% of mesenchymal areas. CK18 stained 100% of epithelial areas. CK19 staining was intense and diffuse in 100% of embryonal samples, but it was weaker in fetal areas (66.66%). AE1 stained epithelial areas in all cases, and it stained 29.41% of mesenchymal areas. AE3 stained 84.61% of embryonal and 60% of fetal components. AE1/AE3 showed stronger staining in embryonal (100%) than in fetal areas (76.92%). Vimentin staining was strong in embryonal (66.66%) and mesenchymal (84.61%) components but weak in fetal areas (8%). Alphafetoprotein was positive in only 20% of fetal and 70% of embryonal areas. Our results support the hypothesis that immunoexpression of HBs follows the stages of normal liver development. Embryonal areas look less differentiated, expressing vimentin and biliary epithelium CKs, whereas fetal areas display a more developed phenotype, similar to that of mature hepatocytes. These data aid in understanding the ontogenesis of HBs and may be used in histopathological diagnosis.