The p53 Pathway Is Frequently Deregulated in Pediatric Burkitt Lymphoma, but over Expression of MDM2 or HDM4 Is Not a Common Mechanism.
Vasiliki Leventaki, Vladimir Rodic, Mona S Jahromi, Jonathan M Downie, Sheryl R Tripp, Michael G Bayerl, Sherrie L Perkins, Phillip Barnette, Joshua D Schiffman, Rodney R Miles. University of Utah, Salt Lake City; ARUP Laboratories, Salt Lake City, UT; Penn State College of Medicine, Hershey, PA
Background: The human homologs of murine double minute 2 (MDM2) and 4 (HDM4) negatively regulate p53. MDM2 over expression and TP53 mutations have been described in pediatric Burkitt lymphoma (pBL), but patterns of HDM4 expression have not been reported. We investigated p53 pathway disruption in pBL by studying HDM4 and MDM2 expression, TP53 mutations, and gene copy number abnormalities.
Design: DNA and RNA were isolated from 30 formalin-fixed, paraffin-embedded diagnostic pBL specimens. Germline DNA from patients' negative staging bone marrows (n=25) served as a pooled normal reference. Tumor and germline DNA were submitted for Molecular Inversion Probe assay (330K Cancer Panel, Affymetrix) and data were analyzed with Nexus Copy Number software (BioDiscovery) for genome-wide copy number, loss of heterozygosity (LOH), and TP53 mutations. MDM2, HDM4, and p53 protein levels were assessed by immunohistochemistry. Relative levels of mRNA for MDM2 and HDM4 [full length (HDM4-FL) and short splicing variant (HDM4-S)] were assayed by q-RT-PCR.
Results: HDM4 protein was expressed in 24/24 tumors with intensity slightly lower than reactive germinal center B-cells in most and equally bright in two cases. HDM4-FL transcripts varied over a 4-fold range among tumor samples, and 3/23 (13%) showed predominance of the HDM4-S (HDM4-S to HDM4-FL ratio>1). MDM2 protein was weakly expressed in 10/28 (35%) and MDM2 mRNA levels showed a 2-4-fold increase (compared to reactive lymph node) with one sample showing a 16-fold increase. HDM4 locus gains were seen in 3/26 patients (12%) while specific amplification or deletion of the MDM2 locus was not seen. TP53 showed deletions (2/26), LOH (3/26), and mutations (5/26). Mutations were seen in 1/2 deleted cases and 2/3 LOH cases, so TP53 genetic changes cumulatively involved 7/26 (27%). p53 protein was expressed in 15/28 (54%) cases, including 6/7 with TP53 genetic alterations. No cases with HDM4 gains showed p53 protein or TP53 genetic alterations. The case with high MDM2 mRNA showed no MDM2, HDM4 or TP53 genetic alterations.
Conclusions: We demonstrate for the first time that HDM4 protein is uniformly expressed in pBL. p53 protein over expression (surrogate for TP53 mutation) was seen in about half of cases including 6/7 with identified TP53 changes. Thus the p53 pathway was deregulated in the majority of our pBL, but significant over expression of HDM4 or MDM2 do not appear to be common mechanisms in our study.
Monday, February 28, 2011 9:30 AM
Poster Session I Stowell-Orbison/Surgical Pathology/Autopsy Awards Poster Session # 185, Monday Morning