Whole genome amplification and molecular genetic analysis of DNA from paraffin-embedded prostate adenocarcinoma tumor tissue

Purpose: Often tissues obtained from prostate adenocarcinoma tumors embedde d in paraffin are heterogeneous in cell type and must be carefully microdis sected to acquire tissue fragments that provide homogeneous aliquots of tum or clones. Such tissue fragments rarely contain sufficient DNA to perform g enomic characterization needed as an early step in localizing relevant onco genes or tumor suppressor genes. We report that PCR using a degenerate olig onucleotide primer (DOP-PCR) can be applied to DNA-samples from microdissec ted paraffin-embedded prostate adenocarcinomas, and this provides sufficien t product for fluorescent allelic imbalance measurements or comparative gen omic hybridization (CGH). Materials and Methods: Samples were selected to be representative of those routinely obtained during prostatectomies,based on typical tumor stages (T2 and T3) and Gleason grades (range 3 + 3 to 4 +5). For DNA analysis without prior DOP-PCR, only large tumors were selected to be sectioned. More than 50 specimens were analyzed. Close comparison of data obtained from analysis of DOP-PCR with those from non-DOP DNA was obtained on a subset 8 samples. To compare the allelic balance of DOP-PCR amplified DNA with that measured for non-DOP DNA, we analyzed allelic ratios on DNA from 5 different tissue samples processed by both microdissection and conventional sectioning. Results: Systematic comparison of allelic imbalance results shows close sim ilarity between DOP-PCR amplified product and non-DOP DNA, indicating that PCR product is a valid representation of the tumor genome. In addition, the difference between allelic balance and imbalance is more distinctive when microdissection followed by DOP-PCR is performed. Performing CGH on product s of DOP-PCR also shows distinctive regional copy number alterations in DNA from microdissected tumor tissue. Conclusion: Either of these procedures allows distinction between benign an d malignant genomes, and also allows independent analysis of genomic altera tions in different portions of tumors. They also may be applied clinically for genomic characterization of small foci that frequently appear in prosta tes of elderly men who are showing no obvious pathological symptoms of aden ocarcinoma.