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Characterization of DNA topoisomerase I in three SN-38 resistant human colon cancer cell lines reveals a new pair of resistance-associated mutations

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Standard

Characterization of DNA topoisomerase I in three SN-38 resistant human colon cancer cell lines reveals a new pair of resistance-associated mutations. / Jensen, Niels Frank; Agama, Keli; Roy, Amit; Smith, David Hersi; Pfister, Thomas D; Rømer, Maria Unni; Zhang, Hong-Liang; Doroshow, James H; Knudsen, Birgitta R; Stenvang, Jan; Brünner, Nils; Pommier, Yves.

I: Journal of Experimental and Clinical Cancer Research (Online), Bind 35, Nr. 1, 56, 31.03.2016.

Publikation: Bidrag til tidsskrift/Konferencebidrag i tidsskrift /Bidrag til avisTidsskriftartikelForskningpeer review

Harvard

Jensen, NF, Agama, K, Roy, A, Smith, DH, Pfister, TD, Rømer, MU, Zhang, H-L, Doroshow, JH, Knudsen, BR, Stenvang, J, Brünner, N & Pommier, Y 2016, 'Characterization of DNA topoisomerase I in three SN-38 resistant human colon cancer cell lines reveals a new pair of resistance-associated mutations', Journal of Experimental and Clinical Cancer Research (Online), bind 35, nr. 1, 56. https://doi.org/10.1186/s13046-016-0335-x

APA

Jensen, N. F., Agama, K., Roy, A., Smith, D. H., Pfister, T. D., Rømer, M. U., ... Pommier, Y. (2016). Characterization of DNA topoisomerase I in three SN-38 resistant human colon cancer cell lines reveals a new pair of resistance-associated mutations. Journal of Experimental and Clinical Cancer Research (Online), 35(1), [56]. https://doi.org/10.1186/s13046-016-0335-x

CBE

Jensen NF, Agama K, Roy A, Smith DH, Pfister TD, Rømer MU, Zhang H-L, Doroshow JH, Knudsen BR, Stenvang J, Brünner N, Pommier Y. 2016. Characterization of DNA topoisomerase I in three SN-38 resistant human colon cancer cell lines reveals a new pair of resistance-associated mutations. Journal of Experimental and Clinical Cancer Research (Online). 35(1). https://doi.org/10.1186/s13046-016-0335-x

MLA

Vancouver

Jensen NF, Agama K, Roy A, Smith DH, Pfister TD, Rømer MU o.a. Characterization of DNA topoisomerase I in three SN-38 resistant human colon cancer cell lines reveals a new pair of resistance-associated mutations. Journal of Experimental and Clinical Cancer Research (Online). 2016 mar 31;35(1). 56. https://doi.org/10.1186/s13046-016-0335-x

Author

Jensen, Niels Frank ; Agama, Keli ; Roy, Amit ; Smith, David Hersi ; Pfister, Thomas D ; Rømer, Maria Unni ; Zhang, Hong-Liang ; Doroshow, James H ; Knudsen, Birgitta R ; Stenvang, Jan ; Brünner, Nils ; Pommier, Yves. / Characterization of DNA topoisomerase I in three SN-38 resistant human colon cancer cell lines reveals a new pair of resistance-associated mutations. I: Journal of Experimental and Clinical Cancer Research (Online). 2016 ; Bind 35, Nr. 1.

Bibtex

@article{639fa58ee4f545d48c5ba52380fa8c04,
title = "Characterization of DNA topoisomerase I in three SN-38 resistant human colon cancer cell lines reveals a new pair of resistance-associated mutations",
abstract = "BACKGROUND: DNA topoisomerase I (Top1) is a DNA unwinding protein and the specific target of the camptothecin class of chemotherapeutic drugs. One of these, irinotecan, acting through its active metabolite SN-38, is used in the treatment of metastatic colorectal cancer. However, resistance to irinotecan represents a major clinical problem. Since molecular alterations in Top1 may result in resistance to irinotecan, we characterized Top1 in three human colon cancer cell lines with acquired resistance to SN-38.METHODS: Three SN-38 resistant (20-67 fold increased resistance) cell lines were generated and compared to wild-type parental cells with regards to: TOP1 gene copy number and gene sequence, Top1 expression (mRNA and protein), Top1 enzymatic activity in the absence and presence of drug, and Top1-DNA cleavage complexes in drug treated cells. TOP1 mutations were validated by PCR using mutant specific primers. Furthermore, cross-resistance to two indenoisoquinoline Top1-targeting drugs (NSC 725776 and NSC 743400) and two Top2-targeting drugs (epirubicin and etoposide) was investigated.RESULTS: Two of three SN-38 resistant cell lines carried TOP1 gene copy number aberrations: A TOP1 gene copy gain and a loss of chromosome 20, respectively. One resistant cell line harbored a pair of yet unreported TOP1 mutations (R364K and G717R) in close proximity to the drug binding site. Mutant TOP1 was expressed at a markedly higher level than wild-type TOP1. None or very small reductions were observed in Top1 expression or Top1 activity in the absence of drug. In all three SN-38 resistant cell lines Top1 activity was maintained in the presence of high concentrations of SN-38. None or only partial cross-resistance were observed for etoposide and epirubicin, respectively. SN-38 resistant cells with wild-type TOP1 remained sensitive to NSC 743400, while cells with mutant TOP1 was fully cross-resistant to both indenoisoquinolines. Top1-DNA cleavage complex formation following drug treatment supported the other findings.CONCLUSIONS: This study adds to the growing knowledge about resistance mechanisms for Top1-targeting chemotherapeutic drugs. Importantly, two yet unreported TOP1 mutations were identified, and it was underlined that cross-resistance to the new indenoisoquinoline drugs depends on the specific underlying molecular mechanism of resistance to SN-38.",
author = "Jensen, {Niels Frank} and Keli Agama and Amit Roy and Smith, {David Hersi} and Pfister, {Thomas D} and R{\o}mer, {Maria Unni} and Hong-Liang Zhang and Doroshow, {James H} and Knudsen, {Birgitta R} and Jan Stenvang and Nils Br{\"u}nner and Yves Pommier",
year = "2016",
month = "3",
day = "31",
doi = "10.1186/s13046-016-0335-x",
language = "English",
volume = "35",
journal = "Journal of Experimental and Clinical Cancer Research (Online)",
issn = "1756-9966",
publisher = "BioMed Central",
number = "1",

}

RIS

TY - JOUR

T1 - Characterization of DNA topoisomerase I in three SN-38 resistant human colon cancer cell lines reveals a new pair of resistance-associated mutations

AU - Jensen, Niels Frank

AU - Agama, Keli

AU - Roy, Amit

AU - Smith, David Hersi

AU - Pfister, Thomas D

AU - Rømer, Maria Unni

AU - Zhang, Hong-Liang

AU - Doroshow, James H

AU - Knudsen, Birgitta R

AU - Stenvang, Jan

AU - Brünner, Nils

AU - Pommier, Yves

PY - 2016/3/31

Y1 - 2016/3/31

N2 - BACKGROUND: DNA topoisomerase I (Top1) is a DNA unwinding protein and the specific target of the camptothecin class of chemotherapeutic drugs. One of these, irinotecan, acting through its active metabolite SN-38, is used in the treatment of metastatic colorectal cancer. However, resistance to irinotecan represents a major clinical problem. Since molecular alterations in Top1 may result in resistance to irinotecan, we characterized Top1 in three human colon cancer cell lines with acquired resistance to SN-38.METHODS: Three SN-38 resistant (20-67 fold increased resistance) cell lines were generated and compared to wild-type parental cells with regards to: TOP1 gene copy number and gene sequence, Top1 expression (mRNA and protein), Top1 enzymatic activity in the absence and presence of drug, and Top1-DNA cleavage complexes in drug treated cells. TOP1 mutations were validated by PCR using mutant specific primers. Furthermore, cross-resistance to two indenoisoquinoline Top1-targeting drugs (NSC 725776 and NSC 743400) and two Top2-targeting drugs (epirubicin and etoposide) was investigated.RESULTS: Two of three SN-38 resistant cell lines carried TOP1 gene copy number aberrations: A TOP1 gene copy gain and a loss of chromosome 20, respectively. One resistant cell line harbored a pair of yet unreported TOP1 mutations (R364K and G717R) in close proximity to the drug binding site. Mutant TOP1 was expressed at a markedly higher level than wild-type TOP1. None or very small reductions were observed in Top1 expression or Top1 activity in the absence of drug. In all three SN-38 resistant cell lines Top1 activity was maintained in the presence of high concentrations of SN-38. None or only partial cross-resistance were observed for etoposide and epirubicin, respectively. SN-38 resistant cells with wild-type TOP1 remained sensitive to NSC 743400, while cells with mutant TOP1 was fully cross-resistant to both indenoisoquinolines. Top1-DNA cleavage complex formation following drug treatment supported the other findings.CONCLUSIONS: This study adds to the growing knowledge about resistance mechanisms for Top1-targeting chemotherapeutic drugs. Importantly, two yet unreported TOP1 mutations were identified, and it was underlined that cross-resistance to the new indenoisoquinoline drugs depends on the specific underlying molecular mechanism of resistance to SN-38.

AB - BACKGROUND: DNA topoisomerase I (Top1) is a DNA unwinding protein and the specific target of the camptothecin class of chemotherapeutic drugs. One of these, irinotecan, acting through its active metabolite SN-38, is used in the treatment of metastatic colorectal cancer. However, resistance to irinotecan represents a major clinical problem. Since molecular alterations in Top1 may result in resistance to irinotecan, we characterized Top1 in three human colon cancer cell lines with acquired resistance to SN-38.METHODS: Three SN-38 resistant (20-67 fold increased resistance) cell lines were generated and compared to wild-type parental cells with regards to: TOP1 gene copy number and gene sequence, Top1 expression (mRNA and protein), Top1 enzymatic activity in the absence and presence of drug, and Top1-DNA cleavage complexes in drug treated cells. TOP1 mutations were validated by PCR using mutant specific primers. Furthermore, cross-resistance to two indenoisoquinoline Top1-targeting drugs (NSC 725776 and NSC 743400) and two Top2-targeting drugs (epirubicin and etoposide) was investigated.RESULTS: Two of three SN-38 resistant cell lines carried TOP1 gene copy number aberrations: A TOP1 gene copy gain and a loss of chromosome 20, respectively. One resistant cell line harbored a pair of yet unreported TOP1 mutations (R364K and G717R) in close proximity to the drug binding site. Mutant TOP1 was expressed at a markedly higher level than wild-type TOP1. None or very small reductions were observed in Top1 expression or Top1 activity in the absence of drug. In all three SN-38 resistant cell lines Top1 activity was maintained in the presence of high concentrations of SN-38. None or only partial cross-resistance were observed for etoposide and epirubicin, respectively. SN-38 resistant cells with wild-type TOP1 remained sensitive to NSC 743400, while cells with mutant TOP1 was fully cross-resistant to both indenoisoquinolines. Top1-DNA cleavage complex formation following drug treatment supported the other findings.CONCLUSIONS: This study adds to the growing knowledge about resistance mechanisms for Top1-targeting chemotherapeutic drugs. Importantly, two yet unreported TOP1 mutations were identified, and it was underlined that cross-resistance to the new indenoisoquinoline drugs depends on the specific underlying molecular mechanism of resistance to SN-38.

U2 - 10.1186/s13046-016-0335-x

DO - 10.1186/s13046-016-0335-x

M3 - Journal article

C2 - 27029323

VL - 35

JO - Journal of Experimental and Clinical Cancer Research (Online)

JF - Journal of Experimental and Clinical Cancer Research (Online)

SN - 1756-9966

IS - 1

M1 - 56

ER -