AIM: To identify the new alternative splicing variants of human CYP2D6 in human extratumoral liver tissue with RT-PCR and sequencing. METHODS: Full length of human CYP2D6 cDNAs was amplificated by reverse transcriptio...AIM: To identify the new alternative splicing variants of human CYP2D6 in human extratumoral liver tissue with RT-PCR and sequencing. METHODS: Full length of human CYP2D6 cDNAs was amplificated by reverse transcription-polymerase chain reaction (RT-PCR) from a human extratumoral liver tissue and cloned into pGEM-T vector. The cDNA was sequenced. Exons from 1 to 4 of human CYP2D6 cDNAs were also amplificated by RT-PCR from extratumoral liver tissues of 17 human hepatocellular carcinomas. Some RT-PCR products were sequenced. Exons 1 to 4 of CYP2D6 gene were amplified by PCR from extratumoral liver tissue DNA. Two PCR products from extratumoral liver tissues expressing skipped mRNA were partially sequenced. RESULTS: One of the CYP2D6 cDNAs had 470 nucleotides from 79 to 548 (3' portion of exons 1 to 5' portion of exon 4), and was skipped. Exons 1 to 4 of CYP2D6 cDNA were assayed with RT-PCR in 17 extratumoral liver tissues. Both wild type and skipped mRNAs were expressed in 4 samples, only wild type mRNA was expressed in 5 samples, and only skipped mRNA was expressed in 8 samples. Two more variants were identified by sequencing the RT-PCR products of exons 1 to 4 of CYP2D6 cDNA. The second variant skipped 411 nucleotides from 175 to 585. This variant was identified in 4 different liver tissues by sequencing the RT-PCR products. We sequenced partially 2 of the PCR products amplified of CYP2D6 exon 1 to exon 4 from extratumoral liver tissue genomic DNA that only expressed skipped mRNA by RT-PCR. No point mutations around exon 1, intron 1, and exon 4, and no deletion in CYP2D6 gene were detected. The third variant was the skipped exon 3, and 153 bp was lost. CONCLUSION: Three new alternative splicing variants of CYP2D6 mRNA have been identified. They may not be caused by gene mutation and may lose CYP2D6 activity and act as a down-regulator of CYP2D6.展开更多
BACKGROUND There are concerns that tamoxifen is less effective in Asian women because of the high prevalence of impaired function cytochrome P4502D6(CYP2D6)polymorphisms.AIM To evaluate how knowledge of CYP2D6 genotyp...BACKGROUND There are concerns that tamoxifen is less effective in Asian women because of the high prevalence of impaired function cytochrome P4502D6(CYP2D6)polymorphisms.AIM To evaluate how knowledge of CYP2D6 genotype impacted the choice of hormonal agent and how CYP2D6 genotype and agent were associated with clinical outcomes.METHODS Eighty-two women were recruited.Seventy-eight completed CYP2D6 genotyping and were categorized into poor,intermediate(IM)and extensive or ultra metabolizer phenotypes.Women with poor metabolizer and IM phenotypes were recommended aromatase inhibitors as the preferred agent.RESULTS More than 70%of the women had an IM phenotype,32%an extensive or ultra metabolizer phenotype,and 0%had a poor metabolizer phenotype.Regardless of genotype,more women opted for aromatase inhibitors.Overall,80%of women completed 5 years of hormonal therapy.Five women developed recurrence,3 contralateral breast cancer,5 died,and 1 was diagnosed with a second primary cancer.Five-year recurrence-free and overall survival were slightly better in women with the extensive or ultra metabolizer phenotype compared to those with the IM phenotype,though not statistically significant[P=0.743,hazard ratio(HR):1.441,95%confidence interval(CI):0.191 to 10.17 and P=0.798,HR:1.327,95%CI:0.172 to 9.915,respectively].Women receiving aromatase inhibitors also appeared to have a better,but also nonsignificant,5-year recurrence-free and overall survival(P=0.253,HR:0.368,95%CI:0.031 to 0.258 and P=0.292,HR:0.252,95%CI:0.005 to 4.951,respectively).CONCLUSION The IM phenotype was highly prevalent but was not associated with clinical outcome.展开更多
基金Natural science foundation project of Heilongjiang Province(D201186)scientific research item of health department of Heilongjiang Province(2012-179)~~
基金Supported by the National Key Basic Research and Development Program of China,No.2002CB512901National Natural Science Foundation of China,No.39770868 and Natural Science Foundation of Zhejiang Province,No.397490
文摘AIM: To identify the new alternative splicing variants of human CYP2D6 in human extratumoral liver tissue with RT-PCR and sequencing. METHODS: Full length of human CYP2D6 cDNAs was amplificated by reverse transcription-polymerase chain reaction (RT-PCR) from a human extratumoral liver tissue and cloned into pGEM-T vector. The cDNA was sequenced. Exons from 1 to 4 of human CYP2D6 cDNAs were also amplificated by RT-PCR from extratumoral liver tissues of 17 human hepatocellular carcinomas. Some RT-PCR products were sequenced. Exons 1 to 4 of CYP2D6 gene were amplified by PCR from extratumoral liver tissue DNA. Two PCR products from extratumoral liver tissues expressing skipped mRNA were partially sequenced. RESULTS: One of the CYP2D6 cDNAs had 470 nucleotides from 79 to 548 (3' portion of exons 1 to 5' portion of exon 4), and was skipped. Exons 1 to 4 of CYP2D6 cDNA were assayed with RT-PCR in 17 extratumoral liver tissues. Both wild type and skipped mRNAs were expressed in 4 samples, only wild type mRNA was expressed in 5 samples, and only skipped mRNA was expressed in 8 samples. Two more variants were identified by sequencing the RT-PCR products of exons 1 to 4 of CYP2D6 cDNA. The second variant skipped 411 nucleotides from 175 to 585. This variant was identified in 4 different liver tissues by sequencing the RT-PCR products. We sequenced partially 2 of the PCR products amplified of CYP2D6 exon 1 to exon 4 from extratumoral liver tissue genomic DNA that only expressed skipped mRNA by RT-PCR. No point mutations around exon 1, intron 1, and exon 4, and no deletion in CYP2D6 gene were detected. The third variant was the skipped exon 3, and 153 bp was lost. CONCLUSION: Three new alternative splicing variants of CYP2D6 mRNA have been identified. They may not be caused by gene mutation and may lose CYP2D6 activity and act as a down-regulator of CYP2D6.
基金Supported by the NHG-KTPH Small Innovative Grants(SIG),No.SIG/11009 and No.SIG/15025.
文摘BACKGROUND There are concerns that tamoxifen is less effective in Asian women because of the high prevalence of impaired function cytochrome P4502D6(CYP2D6)polymorphisms.AIM To evaluate how knowledge of CYP2D6 genotype impacted the choice of hormonal agent and how CYP2D6 genotype and agent were associated with clinical outcomes.METHODS Eighty-two women were recruited.Seventy-eight completed CYP2D6 genotyping and were categorized into poor,intermediate(IM)and extensive or ultra metabolizer phenotypes.Women with poor metabolizer and IM phenotypes were recommended aromatase inhibitors as the preferred agent.RESULTS More than 70%of the women had an IM phenotype,32%an extensive or ultra metabolizer phenotype,and 0%had a poor metabolizer phenotype.Regardless of genotype,more women opted for aromatase inhibitors.Overall,80%of women completed 5 years of hormonal therapy.Five women developed recurrence,3 contralateral breast cancer,5 died,and 1 was diagnosed with a second primary cancer.Five-year recurrence-free and overall survival were slightly better in women with the extensive or ultra metabolizer phenotype compared to those with the IM phenotype,though not statistically significant[P=0.743,hazard ratio(HR):1.441,95%confidence interval(CI):0.191 to 10.17 and P=0.798,HR:1.327,95%CI:0.172 to 9.915,respectively].Women receiving aromatase inhibitors also appeared to have a better,but also nonsignificant,5-year recurrence-free and overall survival(P=0.253,HR:0.368,95%CI:0.031 to 0.258 and P=0.292,HR:0.252,95%CI:0.005 to 4.951,respectively).CONCLUSION The IM phenotype was highly prevalent but was not associated with clinical outcome.
