Genetic Variants of the Copy Number Polymorphic β-Defensin Locus Are Associated with Sporadic Prostate Cancer

Abstract

Background/Aims: Prostate cancer represents the cancer with the highest worldwide prevalence in men. Chromosome 8p23 has shown suggestive genetic linkage to early-onset familial prostate cancer and is frequently deleted in cancer cells of the urogenital tract. Within this locus some β-defensin genes (among them DEFB4, DEFB103, DEFB104) are localized, which are arranged in a gene cluster shown to exhibit an extensive copy number variation in the population. This structural variation considerably hampers genetic studies. In a new approach considering both sequence as well as copy number variations we aimed to compare the defensin locus at 8p23 in prostate cancer patients and controls. Methods: We apply PCR/cloning-based haplotyping and high-throughput copy number determination methods which allow assessment of both individual haplotypes and gene copy numbers not accessible to conventional SNP-based genotyping. Results: We demonstrate association of four common DEFB104 haplotypes with the risk of prostate cancer in two independent patient cohorts. Moreover, we show that high copy numbers (>9) of the defensin gene cluster are significantly underrepresented in both patient samples. Conclusions: Our findings imply a role of the antibacterial defensins in prostate cancerogenesis qualifying distinct gene variants and copy numbers as potential tumor markers.

Analysis of relative gene dosage and expression differences of the paralogs RABL2A and RABL2B by Pyrosequencing

“…Such exploitable sequence variations are allele-discriminating single nucleotide polymorphisms (SNP) or paralog-discriminating sequence variations (PSV). For instance, Deutsch et al (2004) have used Pyrosequencing for a Paralog Ratio Test (PRT) to detect aneuploidies by paralogous sequence quantification (a similar problem as assessed here) and Huse et al (2008) to determine copy numbers of the ß-defensin locus at 8p23. The analytical power of Pyrosequencing analyses has also been shown with the quantification of SNPs across a wide range of allele frequencies in pooled DNA samples (Gruber et al, 2002).…”

“…Pyrosequencing has been widely used for sequencing short stretches of DNA and for the quantitative determination of nucleic acids (Ronaghi et al, 2007). On a genomic level it allows the evaluation of allele dosage/genotypes determined by single nucleotide polymorphisms or structural variations such as copy number variants (Langaee and Ronaghi, 2005;Huse et al, 2008) and it is also used to characterize gene expression (Wittkopp et al, 2008). In Pyrosequencing pyrophosphate is stoichiometrically split off from the deoxynucleoside triphosphates during polymerase reaction and initiates a reaction cascade leading to quantifiable light emission.…”

Analysis of β-defensin and Toll-like receptor gene copy number variation in celiac disease

“…Again, no correlation between copy number and gene function was observed in patients with active psoriasis [30]. Finally, high copy numbers of the gene cluster were shown to be significantly underrepresented in two different prostate cancer patient cohorts, suggesting that genomic CNV in these loci may also be involved in carcinogenesis [31].…”

Comparison of multiplex ligation-dependent probe amplification and real-time PCR accuracy for gene copy number quantification using the β-defensin locus

“…The method of real-time quantitative PCR using LightCycler Relative Quantification Software 1.0 (Roche, Mannheim, Germany) for analysis requires an appropriate calibrator. Genomic DNA sample NA18608 HM , with one copy of target gene per haploid genome as determined by different methods (14,19) and one copy of reference gene per haploid genome, was used as calibrator. The PCR reaction was performed according to the protocol established by our group (13).…”