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The potential role of Alu Y in the development of resistance to SN38 (Irinotecan) or oxaliplatin in colorectal cancer.

Lin X, Stenvang J, Rasmussen MH, Zhu S, Jensen NF, Tarpgaard LS, Yang G, Belling K, Andersen CL, Li J, Bolund L, Brünner N - BMC Genomics (2015)

Bottom Line: Moreover, we extended the RRBS analysis to tumor tissue from 14 patients with colorectal cancer who either did or did not benefit from capecitabine + oxaliplatin treatment.For the clinical samples, we applied a concept of 'DNA methylation entropy' to estimate the diversity of DNA methylation states of the identified resistance phenotype-associated methylation loci observed in the cell line models.Furthermore, we identified an enrichment of Alu Y sequences that likely results from increased integration of new copies of Alu Y sequence in the drug-resistant cell lines.

View Article: PubMed Central - PubMed

Affiliation: Department of Biomedicine, University of Aarhus, the Bartholin Building, DK-8000, Aarhus C, Denmark. xue.lin@biomed.au.dk.

ABSTRACT

Background: Irinotecan (SN38) and oxaliplatin are chemotherapeutic agents used in the treatment of colorectal cancer. However, the frequent development of resistance to these drugs represents a considerable challenge in the clinic. Alus as retrotransposons comprise 11% of the human genome. Genomic toxicity induced by carcinogens or drugs can reactivate Alus by altering DNA methylation. Whether or not reactivation of Alus occurs in SN38 and oxaliplatin resistance remains unknown.

Results: We applied reduced representation bisulfite sequencing (RRBS) to investigate the DNA methylome in SN38 or oxaliplatin resistant colorectal cancer cell line models. Moreover, we extended the RRBS analysis to tumor tissue from 14 patients with colorectal cancer who either did or did not benefit from capecitabine + oxaliplatin treatment. For the clinical samples, we applied a concept of 'DNA methylation entropy' to estimate the diversity of DNA methylation states of the identified resistance phenotype-associated methylation loci observed in the cell line models. We identified different loci being characteristic for the different resistant cell lines. Interestingly, 53% of the identified loci were Alu sequences- especially the Alu Y subfamily. Furthermore, we identified an enrichment of Alu Y sequences that likely results from increased integration of new copies of Alu Y sequence in the drug-resistant cell lines. In the clinical samples, SOX1 and other SOX gene family members were shown to display variable DNA methylation states in their gene regions. The Alu Y sequences showed remarkable variation in DNA methylation states across the clinical samples.

Conclusion: Our findings imply a crucial role of Alu Y in colorectal cancer drug resistance. Our study underscores the complexity of colorectal cancer aggravated by mobility of Alu elements and stresses the importance of personalized strategies, using a systematic and dynamic view, for effective cancer therapy.

No MeSH data available.


Related in: MedlinePlus

The motif of the flanking sequence of Alu elements. A shows the motif of the flanking sequence of the identified Alu elements in all the RRBS samples; B shows the motif of the flanking sequence of the identified Alu elements in set E; C shows the motif of the flanking sequence of all Alu elements in the human genome reference. D shows the motif of the flanking sequence of identified Alu Y subfamily in set E.
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Fig5: The motif of the flanking sequence of Alu elements. A shows the motif of the flanking sequence of the identified Alu elements in all the RRBS samples; B shows the motif of the flanking sequence of the identified Alu elements in set E; C shows the motif of the flanking sequence of all Alu elements in the human genome reference. D shows the motif of the flanking sequence of identified Alu Y subfamily in set E.

Mentions: We applied WebLogo 3.3 [34] to extract flanking sequence (up- and down-stream 20 bp) motif from the Alu elements shared by all the RRBS samples, including both the cell lines and the clinical samples in our study. We identified a symmetric sequence in both flanking sequences of the Alu elements (Figure 5A). Furthermore, we extracted flanking sequence motifs from the identified Alu elements in set E, which presented the Alu elements shared by the 14 clinical samples and the united sets of P, O and S (Figure 5B). To see whether the above Alu elements had unique features (motif sequences differing from the other Alu elements in the human genome), we extracted the flanking sequence motifs for all Alu sequences in the human genome reference (Figure 5C). Interestingly and in general, the flanking sequence of the Alus identified in all the RRBS samples and the Alus identified in set E both showed highly similar motif sequence to that of all Alus in the human genome reference, which is also consistent with the typical Alu target site duplication (TSD) sequence. Alu insertion depends on L1-coded ORP2, and the target site sequence of ORP2 for insertion is typically ‘TTAAAA’ [19]. Our results showed that the identified Alu sequences, either in all the RRBS samples or in set E, typically rely on L1-coded ORP2 for their insertion, which is in accordance with other observations [19]. Additionally, the ranking of the first flanking sequence (on the left flanking sequence) of the Alus in all the RRBS samples from the top to the bottom was A, C, T and G, according to the probability scale. Moreover, according to the probability scale, the ranking of the first flanking sequence of the Alus in set E, from the top to the bottom, was A, C, G and T. By contrast, the ranking of the first flanking sequence of the Alus in the whole human genome was A, T, C and G. We also extracted the flanking sequence motifs for the Alu Y subfamily in set E. The order of ranking the first flanking sequence from the top to the bottom was A, C, T and G, according to the probability scale (Figure 5D). These observations suggest that the additional Alu elements and Alu Y subfamily elements in this study preferably locate in GC-rich region. Since GC-rich regions are also gene-dense regions, our result implies their activity might have an effect on gene function.Figure 5


The potential role of Alu Y in the development of resistance to SN38 (Irinotecan) or oxaliplatin in colorectal cancer.

Lin X, Stenvang J, Rasmussen MH, Zhu S, Jensen NF, Tarpgaard LS, Yang G, Belling K, Andersen CL, Li J, Bolund L, Brünner N - BMC Genomics (2015)

The motif of the flanking sequence of Alu elements. A shows the motif of the flanking sequence of the identified Alu elements in all the RRBS samples; B shows the motif of the flanking sequence of the identified Alu elements in set E; C shows the motif of the flanking sequence of all Alu elements in the human genome reference. D shows the motif of the flanking sequence of identified Alu Y subfamily in set E.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4440512&req=5

Fig5: The motif of the flanking sequence of Alu elements. A shows the motif of the flanking sequence of the identified Alu elements in all the RRBS samples; B shows the motif of the flanking sequence of the identified Alu elements in set E; C shows the motif of the flanking sequence of all Alu elements in the human genome reference. D shows the motif of the flanking sequence of identified Alu Y subfamily in set E.
Mentions: We applied WebLogo 3.3 [34] to extract flanking sequence (up- and down-stream 20 bp) motif from the Alu elements shared by all the RRBS samples, including both the cell lines and the clinical samples in our study. We identified a symmetric sequence in both flanking sequences of the Alu elements (Figure 5A). Furthermore, we extracted flanking sequence motifs from the identified Alu elements in set E, which presented the Alu elements shared by the 14 clinical samples and the united sets of P, O and S (Figure 5B). To see whether the above Alu elements had unique features (motif sequences differing from the other Alu elements in the human genome), we extracted the flanking sequence motifs for all Alu sequences in the human genome reference (Figure 5C). Interestingly and in general, the flanking sequence of the Alus identified in all the RRBS samples and the Alus identified in set E both showed highly similar motif sequence to that of all Alus in the human genome reference, which is also consistent with the typical Alu target site duplication (TSD) sequence. Alu insertion depends on L1-coded ORP2, and the target site sequence of ORP2 for insertion is typically ‘TTAAAA’ [19]. Our results showed that the identified Alu sequences, either in all the RRBS samples or in set E, typically rely on L1-coded ORP2 for their insertion, which is in accordance with other observations [19]. Additionally, the ranking of the first flanking sequence (on the left flanking sequence) of the Alus in all the RRBS samples from the top to the bottom was A, C, T and G, according to the probability scale. Moreover, according to the probability scale, the ranking of the first flanking sequence of the Alus in set E, from the top to the bottom, was A, C, G and T. By contrast, the ranking of the first flanking sequence of the Alus in the whole human genome was A, T, C and G. We also extracted the flanking sequence motifs for the Alu Y subfamily in set E. The order of ranking the first flanking sequence from the top to the bottom was A, C, T and G, according to the probability scale (Figure 5D). These observations suggest that the additional Alu elements and Alu Y subfamily elements in this study preferably locate in GC-rich region. Since GC-rich regions are also gene-dense regions, our result implies their activity might have an effect on gene function.Figure 5

Bottom Line: Moreover, we extended the RRBS analysis to tumor tissue from 14 patients with colorectal cancer who either did or did not benefit from capecitabine + oxaliplatin treatment.For the clinical samples, we applied a concept of 'DNA methylation entropy' to estimate the diversity of DNA methylation states of the identified resistance phenotype-associated methylation loci observed in the cell line models.Furthermore, we identified an enrichment of Alu Y sequences that likely results from increased integration of new copies of Alu Y sequence in the drug-resistant cell lines.

View Article: PubMed Central - PubMed

Affiliation: Department of Biomedicine, University of Aarhus, the Bartholin Building, DK-8000, Aarhus C, Denmark. xue.lin@biomed.au.dk.

ABSTRACT

Background: Irinotecan (SN38) and oxaliplatin are chemotherapeutic agents used in the treatment of colorectal cancer. However, the frequent development of resistance to these drugs represents a considerable challenge in the clinic. Alus as retrotransposons comprise 11% of the human genome. Genomic toxicity induced by carcinogens or drugs can reactivate Alus by altering DNA methylation. Whether or not reactivation of Alus occurs in SN38 and oxaliplatin resistance remains unknown.

Results: We applied reduced representation bisulfite sequencing (RRBS) to investigate the DNA methylome in SN38 or oxaliplatin resistant colorectal cancer cell line models. Moreover, we extended the RRBS analysis to tumor tissue from 14 patients with colorectal cancer who either did or did not benefit from capecitabine + oxaliplatin treatment. For the clinical samples, we applied a concept of 'DNA methylation entropy' to estimate the diversity of DNA methylation states of the identified resistance phenotype-associated methylation loci observed in the cell line models. We identified different loci being characteristic for the different resistant cell lines. Interestingly, 53% of the identified loci were Alu sequences- especially the Alu Y subfamily. Furthermore, we identified an enrichment of Alu Y sequences that likely results from increased integration of new copies of Alu Y sequence in the drug-resistant cell lines. In the clinical samples, SOX1 and other SOX gene family members were shown to display variable DNA methylation states in their gene regions. The Alu Y sequences showed remarkable variation in DNA methylation states across the clinical samples.

Conclusion: Our findings imply a crucial role of Alu Y in colorectal cancer drug resistance. Our study underscores the complexity of colorectal cancer aggravated by mobility of Alu elements and stresses the importance of personalized strategies, using a systematic and dynamic view, for effective cancer therapy.

No MeSH data available.


Related in: MedlinePlus