[1030] Multiplex Ligation-Dependent Probe Amplification (MLPA) Assay – A Novel Approach for Renal Tumor Typing

Maria Tretiakova, Poluru Reddy, Tatjana Antic, Gladell P Paner, Loren Joseph. University of Chicago, Chicago

Background: Distinction between eosinophilic chromophobe renal cell carcinoma (ChRCC) and renal oncocytoma (RO) can be difficult due to significant morphologic and immunophenotypic overlap. RO harbor infrequent loss of chromosomes 1 and Y, whereas ChRCC is characterized by extensive chromosomal alterations and commonly monosomy of one or more chromosomes 1, 2, 6, 10, 17 and 21 with 60-96% frequency. Here we test the utility of the DNA based MLPA in assessing gains and losses of all chromosomes on formalin-fixed paraffin embedded (FFPE) diagnostic tissues in a single tube assay.
Design: MLPA was used to analyze DNA extracted from tumor and matched normal kidney FFPE tissue from 19 patients including 6 classic ChRCC, 7 RO, and 6 cases with hybrid morphology (ChRCC-RO). The SALSA MLPA kit P036-E1 (MRC-Holland) contains a 49 probe mix which targets the subtelomeric regions of each arm of all 23 chromosomes, potentially measuring chromosomal copy number. The pattern of PCR amplicons and their signal strengths were compared (after normalization) to the reference normal samples and control cases with known chromosomal abnormalities.
Results: Five of seven RO (71%) cases showed no chromosomal abnormalities. Two RO cases showed loss of chromosome 20, and one RO had additional loss of chromosome 14. All six classic ChRCC cases showed multiple chromosomal monosomies in various combinations: chromosome 1 (4/6 cases), 2 (3/6 cases), 6 (2/6 cases), 10 (3/6 cases), 13 (2/6 cases), 17 (3/6 cases), X (3/6 cases), and Y (2/3 male patient cases). In addition, we detected loss of chromosomes 3, 5, 8, 12, 21 and 22 with frequency of 17% (1/6 cases each). Among 6 ChRCC-RO cases with overlapping morphologic features 2 cases had no gross chromosomal anomalies, whereas remaining 4 cases had loss of chromosomes 1, 20, 21 and Y in 2 cases, and chromosomes 14 and 17 loss once. Group comparison showed significant difference in chromosomal loss between all studied groups (p<0.05, Fisher exact test).
Conclusions: By MLPA all ChRCC cases showed multiple chromosomal losses with frequent monosomies of chromosomes 1, 2, 6, 10, 13, 17, X and Y, whereas in RO only 29% cases showed single or two chromosome losses. ChRCC-RO group divided with half cases showing chromosomal anomalies similar to ChRCC and half cases with no loss or chromosomal loss similar to RO with monosomy 14 or 20. Our preliminary results show that MLPA could serve as a simple cost-effective method for cytogenetic classification of morphologically challenging renal neoplasms and possibly to identify novel associations in “unclassified” tumors.
Category: Genitourinary (including renal tumors)

Wednesday, March 21, 2012 9:30 AM

Poster Session V # 94, Wednesday Morning


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