Jeffrey S. Dome

Hypomethylation and hypermethylation of DNA in Wilms tumors

We quantitatively analysed hypermethylation at CpG islands in the 5′ ends of 12 genes and one non-CpG island 5′ region (MTHFR) in 31 Wilms tumors. We also determined their global genomic 5-methylcytosine content. Compared with various normal postnatal tissues, ∼40–90% of these pediatric kidney cancers were hypermethylated in four of the genes, MCJ, RASSF1A, TNFRSF12 and CALCA as determined by a quantitative bisulfite-based assay (MethyLight). Interestingly, the non-CpG island 5′ region of MTHFR was less methylated in most tumors relative to the normal tissues. By chromatographic analysis of DNA digested to deoxynucleosides, about 60% of the Wilms tumors were found to be deficient in their overall levels of DNA methylation. We also analysed expression of the three known functional DNA methyltransferase genes. No relationship was observed between global genomic 5-methylcytosine levels and relative amounts of RNA for DNA methyltransferases DNMT1, DNMT3A, and DNMT3B. Importantly, no association was seen between CpG island hypermethylation and global DNA hypomethylation in these cancers. Therefore, the overall genomic hypomethylation frequently observed in cancers is probably not just a response or a prelude to hypermethylation elsewhere in the genome. This suggests that the DNA hypomethylation contributes independently to oncogenesis or tumor progression.

Recent advances in Wilms tumor genetics.

The past decade has witnessed substantial growth in our knowledge of the genes and loci that are altered in Wilms tumor. Although Wilms tumor was one of the original paradigms of Knudsons two-hit model of cancer formation, it has become apparent that several genetic events contribute to Wilms tumorigenesis. Recent research has identified targets and regulators of the first Wilms tumor gene, WT1, has uncovered several candidate genes at the second Wilms tumor locus, WT2, and has identified two familial Wilms tumor loci, FWT1 and FWT2. The recent discovery of activating &bgr;-catenin mutations in some Wilms tumors has also implicated the Wnt signaling pathway in this neoplasm. Recurrent abnormalities of other loci, including 16q, 1p, and 7p, have indicated that these sites may harbor Wilms tumor genes. An enhanced understanding of these and other genetic lesions will provide the foundation for novel targeted Wilms tumor therapies.

Local lymph node involvement does not predict poor outcome in pediatric renal cell carcinoma

Local lymph node involvement in adults with renal cell carcinoma (RCC) is associated with poor outcome. The prognostic significance of local lymph node involvement in children with RCC has not been studied systematically.

Development and functional characterization of human bone marrow mesenchymal cells immortalized by enforced expression of telomerase

Summary. To create immortal mesenchymal cell lines, we transduced primary human bone marrow mesenchymal cells with telomerase reverse transcriptase (TERT). TERT+ mesenchymal cells continued to grow for > 2 years; parallel TERT– cultures underwent senescence after 15 weeks. TERT+ mesenchymal cells did not form foci in soft agar, had a normal karyotype and could differentiate into osteoblasts and chondrocytes. Their capacity to support leukaemic lymphoblasts and normal CD34+ haematopoietic cells was equal to or greater than that of primary cells; 42 TERT+ mesenchymal cell clones varied in their supporting capacity. Immortalized mesenchymal cells offer a promising tool for identifying molecules that regulate human haematopoiesis.

Molecular Characterization of the Pediatric Preclinical Testing Panel

Purpose: Identifying novel therapeutic agents for the treatment of childhood cancers requires preclinical models that recapitulate the molecular characteristics of their respective clinical histotypes. Experimental Design and Results: Here, we have applied Affymetrix HG-U133Plus2 profiling to an expanded panel of models in the Pediatric Preclinical Testing Program. Profiling led to exclusion of two tumor lines that were of mouse origin and five osteosarcoma lines that did not cluster with human or xenograft osteosarcoma samples. We compared expression profiles of the remaining 87 models with profiles from 112 clinical samples representing the same histologies and show that model tumors cluster with the appropriate clinical histotype, once “immunosurveillance” genes (contributed by infiltrating immune cells in clinical samples) are eliminated from the analysis. Analysis of copy number alterations using the Affymetrix 100K single nucleotide polymorphism GeneChip showed that the models have similar copy number alterations to their clinical counterparts. Several consistent copy number changes not reported previously were found (e.g., gain at 22q11.21 that was observed in 5 of 7 glioblastoma samples, loss at 16q22.3 that was observed in 5 of 9 Ewings sarcoma and 4 of 12 rhabdomyosarcoma models, and amplification of 21q22.3 that was observed in 5 of 7 osteosarcoma models). We then asked whether changes in copy number were reflected by coordinate changes in gene expression. We identified 493 copy number–altered genes that are nonrandom and appear to identify histotype-specific programs of genetic alterations. Conclusions: These data indicate that the preclinical models accurately recapitulate expression profiles and genetic alterations common to childhood cancer, supporting their value in drug development.

Improved survival for patients with recurrent Wilms tumor: The experience at St. Jude Children's Research Hospital

Background Reported estimates of survival for patients with recurrent Wilms tumor are 24% to 43%. Because published survival data are more than a decade old and do not reflect advances in therapy, the authors reviewed their experience in treating recurrent Wilms tumor to determine whether the probability of survival has increased. Patients and Methods The authors reviewed the cases of 54 patients with recurrent Wilms tumor who were treated on one of six consecutive clinical trials at St. Jude Childrens Research Hospital between 1969 and 2000. Results Five-year overall survival estimates after relapse were 63.6 ± 15.7% for patients treated during or after 1984 (n = 20) and 20.6 ± 6.5% for patients treated before 1984 (n = 34) (P = 0.002). When the analysis was restricted to patients with high-risk clinical features, 5-year overall survival estimates were 47.6 ± 15.7% for those treated in the modern era (n = 16) and 11.1 ± 5.2% for those treated in the earlier era (n = 25) (P = 0.005). Only three patients received high-dose chemotherapy with autologous stem cell rescue; one survived. No patients with recurrent anaplastic histology disease survived. Conclusions Significant progress has been achieved in the treatment of recurrent favorable-histology Wilms tumor using multimodality salvage regimens with conventional doses of chemotherapy. Novel therapeutic strategies will be necessary to cure patients with recurrent anaplastic Wilms tumor.

Genomic Loss of Tumor Suppressor miRNA-204 Promotes Cancer Cell Migration and Invasion by Activating AKT/mTOR/Rac1 Signaling and Actin Reorganization

Increasing evidence suggests that chromosomal regions containing microRNAs are functionally important in cancers. Here, we show that genomic loci encoding miR-204 are frequently lost in multiple cancers, including ovarian cancers, pediatric renal tumors, and breast cancers. MiR-204 shows drastically reduced expression in several cancers and acts as a potent tumor suppressor, inhibiting tumor metastasis in vivo when systemically delivered. We demonstrated that miR-204 exerts its function by targeting genes involved in tumorigenesis including brain-derived neurotrophic factor (BDNF), a neurotrophin family member which is known to promote tumor angiogenesis and invasiveness. Analysis of primary tumors shows that increased expression of BDNF or its receptor tropomyosin-related kinase B (TrkB) parallel a markedly reduced expression of miR-204. Our results reveal that loss of miR-204 results in BDNF overexpression and subsequent activation of the small GTPase Rac1 and actin reorganization through the AKT/mTOR signaling pathway leading to cancer cell migration and invasion. These results suggest that microdeletion of genomic loci containing miR-204 is directly linked with the deregulation of key oncogenic pathways that provide crucial stimulus for tumor growth and metastasis. Our findings provide a strong rationale for manipulating miR-204 levels therapeutically to suppress tumor metastasis.

Recurrent DGCR8, DROSHA, and SIX Homeodomain Mutations in Favorable Histology Wilms Tumors

We report the most common single-nucleotide substitution/deletion mutations in favorable histology Wilms tumors (FHWTs) to occur within SIX1/2 (7% of 534 tumors) and microRNA processing genes (miRNAPGs) DGCR8 and DROSHA (15% of 534 tumors). Comprehensive analysis of 77 FHWTs indicates that tumors with SIX1/2 and/or miRNAPG mutations show a pre-induction metanephric mesenchyme gene expression pattern and are significantly associated with both perilobar nephrogenic rests and 11p15 imprinting aberrations. Significantly decreased expression of mature Let-7a and the miR-200 family (responsible for mesenchymal-to-epithelial transition) in miRNAPG mutant tumors is associated with an undifferentiated blastemal histology. The combination of SIX and miRNAPG mutations in the same tumor is associated with evidence of RAS activation and a higher rate of relapse and death.

The feasibility and outcome of nephron-sparing surgery for children with bilateral Wilms tumor. The St Jude Children's Research Hospital experience: 1999-2006.

Approximately 5% of children with Wilms tumor present with bilateral disease. The treatment challenge is to achieve a high cure rate while maintaining adequate long‐term renal function. The authors of this report assessed the feasibility and outcome of nephron‐sparing surgery in patients with bilateral Wilms tumor who were treated at a single institution.

Telomerase Expression Predicts Unfavorable Outcome in Osteosarcoma

PURPOSE Osteosarcoma is distinct from most cancers in that the majority of osteosarcomas lack telomerase expression and use the alternative lengthening of telomeres (ALT) mechanism to maintain telomeres. Laboratory studies suggest that compared with ALT, telomerase expression is associated with increased tumor aggressiveness. We evaluated the clinical significance of telomerase expression in human osteosarcoma. PATIENTS AND METHODS Fifty-six osteosarcomas from 51 patients treated at St Jude Childrens Research Hospital between 1982 and 2003 were evaluated for telomerase enzyme activity, mRNA expression of the catalytic component of telomerase (TERT), and presence of the ALT pathway. RESULTS Outcome analysis was based on TERT mRNA expression in the primary tumor samples from 44 patients. Fourteen primary tumors expressed TERT mRNA (32%; eight TERT only, six TERT and ALT) and 30 did not express TERT mRNA (68%; 29 ALT, one no ALT). Progression-free survival (PFS) was inferior in the TERT-positive group compared with the TERT-negative group (3-year estimates, 21.4% +/- 9.5% v 63.7% +/- 11.1%; P =.014). Likewise, overall survival was inferior in the TERT-positive group compared with the TERT-negative group (3-year estimates, 42.9% +/- 12.2% v 70.0% +/- 9.9%; P =.031). Among 31 patients with nonmetastatic disease at diagnosis, PFS was lower in the TERT-positive group compared with the TERT-negative group (3-year estimates, 33.3% +/- 13.6% v 72.0% +/- 11.5%; P =.092). CONCLUSION Telomerase expression in primary tumor samples is associated with decreased PFS and OS in patients with osteosarcoma. Additional studies are warranted to better define the clinical utility of this molecular marker.