Jun Sugimura

Robust Classification of Renal Cell Carcinoma Based on Gene Expression Data and Predicted Cytogenetic Profiles

Renal cell carcinoma (RCC) is a heterogeneous disease that includes several histologically distinct subtypes. The most common RCC subtypes are clear cell, papillary, and chromophobe, and recent gene expression profiling studies suggest that classification of RCC based on transcriptional signatures could be beneficial. Traditionally, however, patterns of chromosomal alterations have been used to assist in the molecular classification of RCC. The purpose of this study was to determine whether it was possible to develop a classification model for the three major RCC subtypes that utilizes gene expression profiles as the bases for both molecular genetic and cytogenetic classification. Gene expression profiles were first used to build an expression-based RCC classifier. The RCC gene expression profiles were then examined for the presence of regional gene expression biases. Regional expression biases are genetic intervals that contain a disproportionate number of genes that are coordinately up- or down-regulated. The presence of a regional gene expression bias often indicates the presence of a chromosomal abnormality. In this study, we demonstrate an expression-based classifier can distinguish between the three most common RCC subtypes in 99% of cases (n = 73). We also demonstrate that detection of regional expression biases accurately identifies cytogenetic features common to RCC. Additionally, the in silico-derived cytogenetic profiles could be used to classify 81% of cases. Taken together, these data demonstrate that it is possible to construct a robust classification model for RCC using both transcriptional and cytogenetic features derived from a gene expression profile.

The t(1;3) breakpoint-spanning genes LSAMP and NORE1 are involved in clear cell renal cell carcinomas

By positional cloning, we identified two breakpoint-spanning genes in a familial clear cell renal cell carcinoma (CCRCC)-associated t(1;3)(q32.1;q13.3): LSAMP and NORE1 (RASSF1 homolog). Both genes are downregulated in 9 of 9 RCC cell lines. While the NORE1A promoter predominantly presents partial methylation in 6 of the cell lines and 17/53 (32%) primary tumors, the LSAMP promoter is completely methylated in 5 of 9 cell lines and in 14/53 (26%) sporadic and 4 familial CCRCCs. Expression of LSAMP and NORE1A proteins in CCRCC cell lines inhibited cell proliferation. These characteristics indicate that LSAMP and NORE1A may represent new candidate tumor suppressors for CCRCC.

Classification of Renal Neoplasms Based on Molecular Signatures

PURPOSE Gene expression microarray studies have demonstrated distinct molecular signatures for different types of renal neoplasms based on overall gene expression patterns. However, in most of these studies the investigators used renal tumors with defined histology. We analyzed a test set of renal tumors in double-blind fashion using recently established molecular profiles of renal tumors as benchmarks. MATERIALS AND METHODS A total of 16 consecutive nephrectomies performed for neoplasms at a single urological service were subjected to gene expression profiling using cDNA chips containing 21,632 genes. Analysis was clustered with our previously established molecular profiles of 91 histologically defined kidney neoplasms and comparative genomic microarray analysis while blinded to tumor histology and clinical information. RESULTS With molecular analysis 9, 4, 2 and 1 tumors were classified as clear cell, papillary RCC, chromophobe RCC, and renal oncocytoma, respectively. Histopathological evaluation was concordant in 14 tumors. One of the 2 tumors with a discrepancy between molecular and pathological diagnoses was composed of oncocytoma and high grade clear cell RCC, and the other was chromophobe RCC that histologically mimicked papillary RCC. CONCLUSIONS We report the feasibility of the molecular diagnosis and classification of unknown renal neoplasms. Molecular diagnosis appears to be reliable and comparable to the standard of urological pathology. This molecular method may be a potentially useful test for establishing an accurate diagnosis that can impact clinical management.

Genomic expression and single-nucleotide polymorphism profiling discriminates chromophobe renal cell carcinoma and oncocytoma

BackgroundChromophobe renal cell carcinoma (chRCC) and renal oncocytoma are two distinct but closely related entities with strong morphologic and genetic similarities. While chRCC is a malignant tumor, oncocytoma is usually regarded as a benign entity. The overlapping characteristics are best explained by a common cellular origin, and the biologic differences between chRCC and oncocytoma are therefore of considerable interest in terms of carcinogenesis, diagnosis and clinical management. Previous studies have been relatively limited in terms of examining the differences between oncocytoma and chromophobe RCC.MethodsGene expression profiling using the Affymetrix HGU133Plus2 platform was applied on chRCC (n = 15) and oncocytoma specimens (n = 15). Supervised analysis was applied to identify a discriminatory gene signature, as well as differentially expressed genes. High throughput single-nucleotide polymorphism (SNP) genotyping was performed on independent samples (n = 14) using Affymetrix GeneChip Mapping 100 K arrays to assess correlation between expression and gene copy number. Immunohistochemical validation was performed in an independent set of tumors.ResultsA novel 14 probe-set signature was developed to classify the tumors internally with 93% accuracy, and this was successfully validated on an external data-set with 94% accuracy. Pathway analysis highlighted clinically relevant dysregulated pathways of c-erbB2 and mammalian target of rapamycin (mTOR) signaling in chRCC, but no significant differences in p-AKT or extracellular HER2 expression was identified on immunohistochemistry. Loss of chromosome 1p, reflected in both cytogenetic and expression analysis, is common to both entities, implying this may be an early event in histogenesis. Multiple regional areas of cytogenetic alterations and corresponding expression biases differentiating the two entities were identified. Parafibromin, aquaporin 6, and synaptogyrin 3 were novel immunohistochemical markers effectively discriminating the two pathologic entities.ConclusionsGene expression profiles, high-throughput SNP genotyping, and pathway analysis effectively distinguish chRCC from oncocytoma. We have generated a novel transcript predictor that is able to discriminate between the two entities accurately, and which has been validated both in an internal and an independent data-set, implying generalizability. A cytogenetic alteration, loss of chromosome 1p, common to renal oncocytoma and chRCC has been identified, providing the opportunities for identifying novel tumor suppressor genes and we have identified a series of immunohistochemical markers that are clinically useful in discriminating chRCC and oncocytoma.

Gene Expression Profiling of Early- and Late-Relapse Nonseminomatous Germ Cell Tumor and Primitive Neuroectodermal Tumor of the Testis

Purpose: To better understand the molecular mechanisms that underlay the development and progression of nonseminomatous germ cell tumor of testis (NSGCTT) as well as malignant transformation of teratoma and primitive neuroectodermal tumor (PNET). Experimental Design: We studied the gene expression profiles of 17 retroperitoneal NSGCTTs (10 yolk sac tumors, 3 embryonal carcinomas, 4 teratomas) and 2 PNETs obtained from patients with two clinical outcomes. Tissue samples were obtained from the Indiana University. One group of NSGCTT and PNET patients developed metastases within 2 years (early-relapse) of initial successful treatment, and the other group developed metastases after 2 years (late-relapse). Gene expression in these groups of patients was quantified using cDNA microarrays and real-time relative quantitative PCR. Results: We demonstrate that the gene expression profiles of these tumors correlate with histological type. In addition, we identify type-specific genes that may serve as novel diagnostic markers. We also identify a gene set that can distinguish between early-relapse and late-relapse yolk sac tumors. The expression differences of these genes may underlie the differences in clinical outcome and drug response of these tumors. Conclusion: This is the first study that used gene expression profiling to examine the molecular characteristics of the NSGCTTs and drug response in early- and late-relapse tumors. These results suggest that two molecularly distinct forms of NSGCTTs exist and that the integration of expression profile data with clinical parameters could enhance the diagnosis and prognosis of NSGCTTs. More importantly, the identified genes provide insight into the molecular mechanisms of aggressive NSGCTTs and suggest intervention strategies.

Overexpression of glutathione s-transferase alpha in clear cell renal cell carcinoma.

To determine its diagnostic value, we evaluated glutathione S-transferase alpha (GST-alpha) expression in a large number of renal cell carcinomas (RCCs). GST-alpha messenger RNA (mRNA) levels from 70 renal neoplasms were analyzed with complementary DNA (cDNA) microarray chips containing 21,632 cDNA clones. Furthermore, 348 primary renal tumors and 24 metastatic RCCs were subjected to immunohistochemical analysis with a GST-alpha-specific antibody. GST-alpha mRNA was elevated significantly (11.4-fold) in a majority of clear cell RCCs (28/43 [65.1%]; 28/39 [71.8%] with adjustments for informative spots) compared with other kidney tumors (1/27 [3.7%]). Strong and diffuse GST-alpha immunoreactivity was demonstrated in a majority of clear cell (166/202 [82.2%]; mean intensity, 2.41) and metastatic clear cell RCCs (17/24 [70.8%]; mean intensity, 2.62). Other renal tumor types did not exhibit significant GST-alpha immunoreactivity, confirming mRNA results. Through cDNA microarrays and immunohistochemical analysis, we demonstrated GST-alpha as a biomarker for clear cell RCCs.

Gene Expression Profiling of Renal Cell Carcinoma and Its Implications in Diagnosis, Prognosis, and Therapeutics

Renal cell carcinoma (RCC) is the 10th most common cancer in the United States. It is a histologically heterogeneous disease with various histologic types being characterized by distinct genetic alterations. This chapter reviews advances in the gene expression profiling of RCC and discusses their clinical implications. Data are promising, and many more RCC-related microarray studies are currently underway or in planning. Undoubtedly these data will have an impact on the diagnosis, prognosis, and treatment of RCCs in the future. Finally, this chapter discusses what additional studies should be performed to help uncover the molecular mechanisms of RCC and to bring this new knowledge into use in the clinical arena.

Overexpression of Glutathione S-Transferase α in Clear Cell Renal Cell Carcinoma

To determine its diagnostic value, we evaluated glutathione S-transferase α (GST-α) expression in a large number of renal cell carcinomas (RCCs). GST-α messenger RNA (mRNA) levels from 70 renal neoplasms were analyzed with complementary DNA (cDNA) microarray chips containing 21,632 cDNA clones. Furthermore, 348 primary renal tumors and 24 metastatic RCCs were subjected to immunohisto-chemical analysis with a GST-α–specific antibody. GST-α mRNA was elevated significantly (11.4-fold) in a majority of clear cell RCCs (28/43 [65.1%]; 28/39 [71.8%] with adjustments for informative spots) compared with other kidney tumors (1/27 [3.7%]). Strong and diffuse GST-α immunoreactivity was demonstrated in a majority of clear cell (166/202 [82.2%]; mean intensity, 2.41) and metastatic clear cell RCCs (17/24 [70.8%]; mean intensity, 2.62). Other renal tumor types did not exhibit significant GST-α immunoreactivity, confirming mRNA results. Through cDNA microarrays and immunohistochemical analysis, we demonstrated GST-α as a biomarker for clear cell RCCs.

Renal small cell carcinoma (neuroendocrine carcinoma) without features of transitional cell carcinoma

Seventeen cases of renal small cell carcinoma have been reported in the literature. Approximately half of the reported cases show combined features of transitional cell carcinoma. Presented herein is a case of renal small cell carcinoma In a 37‐year‐old Japanese male who had been treated for 10 years with famotidine for duodenal ulcer. He suffered from sudden‐onset chest pain at presentation and myxoma of the right atrium was suspected. He was treated by atrlotomy and a tumor was removed from the right atrium and pulmonary artery. Histological examination, however, revealed It to be small cell carcinoma. Accordingly, a radical operation was performed for the removal of a tumor found in the right kidney. Histological examination of the tumor confirmed the presence of renal small cell carcinoma without any features of transitional cell carcinoma. It is reported that long‐term administration of an histamine 2 (H2) receptor antagonist may produce carcinold tumors in rodents and enterochromaffin‐like cell hyperplasia In humans. The possible relationship between neuroendocrine carcinoma and H2 receptor antagonist therapy is discussed.

Telomerase activity in human renal cell carcinoma

Purpose : Telomerase activity has been detected in a wide variety of human tumors. The present study evaluated telomerase activity in association with the acquisition of renal cell carcinoma (RCC).