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Polymorphisms of glutathione-S-transferase M1, T1, P1 and the risk of prostate cancer: a case-control study.

Sivonová M, Waczulíková I, Dobrota D, Matáková T, Hatok J, Racay P, Kliment J - J. Exp. Clin. Cancer Res. (2009)

Bottom Line: Analysis for the GST gene polymorphisms was performed by PCR and PCR-RFLP.We found that the GST frequencies are not significantly different from those estimated in a European multicentre study or from the results published by another group in Slovakia.Understanding the contribution of GST gene polymorphisms and their interactions with other relevant factors may improve screening diagnostic assays for prostate cancer.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Medical Biochemistry, Comenius University, Jessenius School of Medicine, Martin, Slovakia. sivonova@jfmed.uniba.sk

ABSTRACT

Background: It has been suggested that polymorphisms in glutathione-S-transferases (GST) could predispose to prostate cancer through a heritable deficiency in detoxification pathways for environmental carcinogens. Yet, studies linking GST polymorphism and prostate cancer have so far failed to unambiguously establish this relation in patients. A retrospective study on healthy, unrelated subjects was conducted in order to estimate the population GST genotype frequencies in the Slovak population of men and compare our results with already published data (GSEC project-Genetic Susceptibility to Environmental Carcinogens). A further aim of the study was to evaluate polymorphisms in GST also in patients with prostate cancer in order to compare the evaluated proportions with those found in the control subjects.

Methods: We determined the GST genotypes in 228 healthy, unrelated subjects who attended regular prostate cancer screening between May 2005 and June 2007 and in 129 histologically verified prostate cancer patients. Analysis for the GST gene polymorphisms was performed by PCR and PCR-RFLP.

Results: We found that the GST frequencies are not significantly different from those estimated in a European multicentre study or from the results published by another group in Slovakia. Our results suggest that Val/Val genotype of GSTP1 gene could modulate the risk of prostate cancer, even if this association did not reach statistical significance. We did not observe significantly different crude rates of the GSTM1 and GSTT1 genotypes in the men diagnosed with prostate cancer and those in the control group.

Conclusion: Understanding the contribution of GST gene polymorphisms and their interactions with other relevant factors may improve screening diagnostic assays for prostate cancer. We therefore discuss issues of study feasibility, study design, and statistical power, which should be taken into account in planning further trials.

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Cleavage of 442 bp PCR products of GSTP1 gene by the Alw26I restriction endonuclease. Ethidium bromide-stained electrophoresed representative PCR-RFLP products samples: 100 bp ladder (lane L), Ile/Ile allele (lanes 2, 3, 5, 6); Ile/Val allele (lanes 1, 7, 8, 9) and Val/Val allele (lane 4).
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Figure 2: Cleavage of 442 bp PCR products of GSTP1 gene by the Alw26I restriction endonuclease. Ethidium bromide-stained electrophoresed representative PCR-RFLP products samples: 100 bp ladder (lane L), Ile/Ile allele (lanes 2, 3, 5, 6); Ile/Val allele (lanes 1, 7, 8, 9) and Val/Val allele (lane 4).

Mentions: The GSTP1 Ile105Val substitution was detected using the PCR-RFLP approach as the substitution by guanine introduced restriction site that can be recognized by an endonuclease Alw26I. PCR reactions were performed in a total volume of 25 μl of solution containing 10 × PCR buffer (16.6 mmol/l (NH4)2SO4, 20.0 mmol/l MgCl2, pH 8.8, 1.2 μl DMSO, 1.2 μl DTT), 200 μmol/l deoxynucleoside triphosphates, 1 U of Taq DNA polymerase, 100 ng of genomic DNA and 25 pmol of GSTP1 primers (forward 5'-GTA GTT TGC CCA AGG TCA AG-3' and reverse 5'-AGC CAC CTG AGG GGT AAG-3', GenBank accession no. NM_000852). The reaction started for 3 min at 94°C, followed by 5 cycles of PCR (cycle 1: 94°C for 15 s, 64°C for 30 s, and 72°C for 1 min) during which the annealing temperature decreased by 1°C for each cycle. This step was followed by 30 cycles of denaturation (for 15 s at 94°C), annealing (for 30 s at 59°C), and extension (for 1 min at 72°C). A final polymerization step (for 5 min at 72°C) was carried out to complete the elongation process and yield a 442-bp fragment. A negative control (PCR without template) was included in each set of PCR reactions. Each PCR product (10 μl) was digested for 4 hours with the restriction enzyme Alw26I (5 U) and electrophoresed on ethidium-bromide-stained 1.5% agarose gel. The presence of the Ile/Ile allele was detected by 329-, and 113-bp fragments, whereas the Val/Val allele was confirmed by 216-, and 113-bp fragments. The heterozygote Ile/Val allele was characterized by fragments consisting of 329, 216, and 113 bp (Fig. 2) [7].


Polymorphisms of glutathione-S-transferase M1, T1, P1 and the risk of prostate cancer: a case-control study.

Sivonová M, Waczulíková I, Dobrota D, Matáková T, Hatok J, Racay P, Kliment J - J. Exp. Clin. Cancer Res. (2009)

Cleavage of 442 bp PCR products of GSTP1 gene by the Alw26I restriction endonuclease. Ethidium bromide-stained electrophoresed representative PCR-RFLP products samples: 100 bp ladder (lane L), Ile/Ile allele (lanes 2, 3, 5, 6); Ile/Val allele (lanes 1, 7, 8, 9) and Val/Val allele (lane 4).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2654432&req=5

Figure 2: Cleavage of 442 bp PCR products of GSTP1 gene by the Alw26I restriction endonuclease. Ethidium bromide-stained electrophoresed representative PCR-RFLP products samples: 100 bp ladder (lane L), Ile/Ile allele (lanes 2, 3, 5, 6); Ile/Val allele (lanes 1, 7, 8, 9) and Val/Val allele (lane 4).
Mentions: The GSTP1 Ile105Val substitution was detected using the PCR-RFLP approach as the substitution by guanine introduced restriction site that can be recognized by an endonuclease Alw26I. PCR reactions were performed in a total volume of 25 μl of solution containing 10 × PCR buffer (16.6 mmol/l (NH4)2SO4, 20.0 mmol/l MgCl2, pH 8.8, 1.2 μl DMSO, 1.2 μl DTT), 200 μmol/l deoxynucleoside triphosphates, 1 U of Taq DNA polymerase, 100 ng of genomic DNA and 25 pmol of GSTP1 primers (forward 5'-GTA GTT TGC CCA AGG TCA AG-3' and reverse 5'-AGC CAC CTG AGG GGT AAG-3', GenBank accession no. NM_000852). The reaction started for 3 min at 94°C, followed by 5 cycles of PCR (cycle 1: 94°C for 15 s, 64°C for 30 s, and 72°C for 1 min) during which the annealing temperature decreased by 1°C for each cycle. This step was followed by 30 cycles of denaturation (for 15 s at 94°C), annealing (for 30 s at 59°C), and extension (for 1 min at 72°C). A final polymerization step (for 5 min at 72°C) was carried out to complete the elongation process and yield a 442-bp fragment. A negative control (PCR without template) was included in each set of PCR reactions. Each PCR product (10 μl) was digested for 4 hours with the restriction enzyme Alw26I (5 U) and electrophoresed on ethidium-bromide-stained 1.5% agarose gel. The presence of the Ile/Ile allele was detected by 329-, and 113-bp fragments, whereas the Val/Val allele was confirmed by 216-, and 113-bp fragments. The heterozygote Ile/Val allele was characterized by fragments consisting of 329, 216, and 113 bp (Fig. 2) [7].

Bottom Line: Analysis for the GST gene polymorphisms was performed by PCR and PCR-RFLP.We found that the GST frequencies are not significantly different from those estimated in a European multicentre study or from the results published by another group in Slovakia.Understanding the contribution of GST gene polymorphisms and their interactions with other relevant factors may improve screening diagnostic assays for prostate cancer.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Medical Biochemistry, Comenius University, Jessenius School of Medicine, Martin, Slovakia. sivonova@jfmed.uniba.sk

ABSTRACT

Background: It has been suggested that polymorphisms in glutathione-S-transferases (GST) could predispose to prostate cancer through a heritable deficiency in detoxification pathways for environmental carcinogens. Yet, studies linking GST polymorphism and prostate cancer have so far failed to unambiguously establish this relation in patients. A retrospective study on healthy, unrelated subjects was conducted in order to estimate the population GST genotype frequencies in the Slovak population of men and compare our results with already published data (GSEC project-Genetic Susceptibility to Environmental Carcinogens). A further aim of the study was to evaluate polymorphisms in GST also in patients with prostate cancer in order to compare the evaluated proportions with those found in the control subjects.

Methods: We determined the GST genotypes in 228 healthy, unrelated subjects who attended regular prostate cancer screening between May 2005 and June 2007 and in 129 histologically verified prostate cancer patients. Analysis for the GST gene polymorphisms was performed by PCR and PCR-RFLP.

Results: We found that the GST frequencies are not significantly different from those estimated in a European multicentre study or from the results published by another group in Slovakia. Our results suggest that Val/Val genotype of GSTP1 gene could modulate the risk of prostate cancer, even if this association did not reach statistical significance. We did not observe significantly different crude rates of the GSTM1 and GSTT1 genotypes in the men diagnosed with prostate cancer and those in the control group.

Conclusion: Understanding the contribution of GST gene polymorphisms and their interactions with other relevant factors may improve screening diagnostic assays for prostate cancer. We therefore discuss issues of study feasibility, study design, and statistical power, which should be taken into account in planning further trials.

Show MeSH
Related in: MedlinePlus