This is a preprint, a preliminary version of a manuscript that has not completed peer review at a journal. Research Square does not conduct peer review prior to posting preprints. The posting of a preprint on this server should not be interpreted as an endorsement of its validity or suitability for dissemination as established information or for guiding clinical practice.
Research
Zhenfeng Duan
University of California Los Angeles David Geffen School of Medicine
Corresponding Author
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Background: Although ATR has an established role in DNA damage response in various cancers, its clinical and prognostic significance in ovarian cancer remains largely unknown. The aims of this study are to assess the expression, function and clinical prognostic relationship of ATR, p-ATR in ovarian cancer.
Methods: We confirmed ATR and p-ATR expressions by immunohistochemistry in a unique ovarian cancer tissue microarray constructed of paired primary, recurrent and metastatic tumor tissues from 26 individual patients. ATR specific siRNA and ATR inhibitor VE-822 were applied to determine the effect of ATR inhibition on ovarian cancer cell proliferation, apoptosis, and DNA damage. ATR expression and the associated proteins of the ATR/Chk1 pathway in ovarian cancer cell lines were evaluated by Western blotting. The clonogenicity was also examined using clonogenic assays. A 3D cell culture model was performed to mimic the in vivo ovarian cancer environment to further validate the effect of ATR inhibition on ovarian cancer cells.
Results: We show recurrent ovarian cancer tissues express higher levels of ATR and p-ATR than their patient-matched primary tumor counterparts. Additionally, higher expression of p-ATR correlates with decreased survival in ovarian cancer patients. Treatment of ovarian cancer cells with ATR specific siRNA or ATR inhibitor VE-822 led to significant apoptosis and inhibition of cellular proliferation, with reduced phosphorylation of Chk1 (p-Chk1), Cdc25c (p-Cdc25c), Cdc2 (p-Cdc2), and increased expression of cleaved PARP and γH2AX. Inhibition of ATR also suppressed clonogenicity and spheroid growth of ovarian cancer cells.
Conclusion: Our results support the ATR and p-ATR pathway as a prognostic biomarker, and targeting the ATR machinery is an emerging therapeutic approach in the treatment of ovarian cancer.
Keywords
ATR, p-ATR, Ovarian cancer, Tissue microarray, Prognostic marker, Therapeutic target
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This is a preprint, a preliminary version of a manuscript that has not completed peer review at a journal. Research Square does not conduct peer review prior to posting preprints. The posting of a preprint on this server should not be interpreted as an endorsement of its validity or suitability for dissemination as established information or for guiding clinical practice.
Research
Zhenfeng Duan
University of California Los Angeles David Geffen School of Medicine
Corresponding Author
Badges
Prescreen
This manuscript passed our Prescreen assessment
Complete author information
Appropriate declaration statements
Potential risks to human health
Prescreen
History
CURRENT STATUS: Posted
View the published article at Therapeutic Advances in Medical Oncology.
Version 1
Posted 16 Apr, 2020
No community comments so far
Metrics
Comments: 0
PDF Downloads: ...
HTML Views: ...
License:
This work is licensed under a CC BY 4.0 License. Read Full License
View the published article at Therapeutic Advances in Medical Oncology.
Background: Although ATR has an established role in DNA damage response in various cancers, its clinical and prognostic significance in ovarian cancer remains largely unknown. The aims of this study are to assess the expression, function and clinical prognostic relationship of ATR, p-ATR in ovarian cancer.
Methods: We confirmed ATR and p-ATR expressions by immunohistochemistry in a unique ovarian cancer tissue microarray constructed of paired primary, recurrent and metastatic tumor tissues from 26 individual patients. ATR specific siRNA and ATR inhibitor VE-822 were applied to determine the effect of ATR inhibition on ovarian cancer cell proliferation, apoptosis, and DNA damage. ATR expression and the associated proteins of the ATR/Chk1 pathway in ovarian cancer cell lines were evaluated by Western blotting. The clonogenicity was also examined using clonogenic assays. A 3D cell culture model was performed to mimic the in vivo ovarian cancer environment to further validate the effect of ATR inhibition on ovarian cancer cells.
Results: We show recurrent ovarian cancer tissues express higher levels of ATR and p-ATR than their patient-matched primary tumor counterparts. Additionally, higher expression of p-ATR correlates with decreased survival in ovarian cancer patients. Treatment of ovarian cancer cells with ATR specific siRNA or ATR inhibitor VE-822 led to significant apoptosis and inhibition of cellular proliferation, with reduced phosphorylation of Chk1 (p-Chk1), Cdc25c (p-Cdc25c), Cdc2 (p-Cdc2), and increased expression of cleaved PARP and γH2AX. Inhibition of ATR also suppressed clonogenicity and spheroid growth of ovarian cancer cells.
Conclusion: Our results support the ATR and p-ATR pathway as a prognostic biomarker, and targeting the ATR machinery is an emerging therapeutic approach in the treatment of ovarian cancer.
Figure 2
Figure 3
Figure 6
Figure 8
Figure 9
Figure 11
This is a list of supplementary files associated with this preprint. Click to download.
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