Advertisement

Breast Cancer Research and Treatment

, Volume 150, Issue 2, pp 299–307 | Cite as

PIK3CA mutations in serum DNA are predictive of recurrence in primary breast cancer patients

  • Chiya Oshiro
  • Naofumi KagaraEmail author
  • Yasuto Naoi
  • Masafumi Shimoda
  • Atsushi Shimomura
  • Naomi Maruyama
  • Kenzo Shimazu
  • Seung Jin Kim
  • Shinzaburo Noguchi
Preclinical Study

Abstract

We attempted to develop a highly sensitive and specific method for the detection of circulating tumor DNA (ctDNA) using a digital PCR (dPCR) assay for PIK3CA mutations (i.e., H1047R, E545K, and E542K) in primary breast cancer patients. The sensitivity of the dPCR assay for the mutant alleles was examined using cell lines with PIK3CA mutations and proved to be 0.01 %. Serum samples were collected pre-operatively from 313 stage I–III breast cancer patients, of whom 110 were found to have PIK3CA mutant tumors. The serum samples from these patients with PIK3CA mutant tumors were subjected to the dPCR assay, and 25 (22.7 %) were found to be positive. No PIK3CA mutant ctDNA was detected in the serum samples of 50 healthy women and 30 breast cancer patients with PIK3CA non-mutant tumors. The patients with PIK3CA mutant ctDNA were dichotomized into mutant ctDNA-high (ctDNAhigh) and ctDNA-low (ctDNAlow) groups based on the median. The ctDNAhigh patients exhibited significantly shorter recurrence-free survival (RFS; P = 0.0002) and overall survival rates (OS; P = 0.0048) compared to those exhibited by the combined ctDNAlow patient and ctDNA-free patient group. Multivariate analysis revealed that ctDNAhigh status significantly predicted poor RFS and OS and did so independently of conventional histological parameters. These results suggest that dPCR is a highly sensitive and specific method for the detection of PIK3CA mutant ctDNA and that ctDNAhigh but not ctDNAlow status is a significant and independent prognostic factor for primary breast cancer patients.

Keywords

Breast cancer PIK3CA mutation ctDNA dPCR Prognosis 

Notes

Acknowledgments

This work was supported in part by Grants-in-Aid from the Knowledge Cluster Initiative of the Ministry of Education, Culture, Sports, Science and Technology, Japan.

Conflict of interest

The authors of this study have no conflicts of interest and no financial disclosures to make.

Supplementary material

10549_2015_3322_MOESM1_ESM.pdf (346 kb)
Supplementary material 1 (PDF 346 kb)
10549_2015_3322_MOESM2_ESM.pdf (77 kb)
Supplementary material 2 (PDF 77 kb)

References

  1. 1.
    Crowley E, Di Nicolantonio F, Loupakis F, Bardelli A (2013) Liquid biopsy: monitoring cancer-genetics in the blood. Nat Rev Clin Oncol 10(8):472–484. doi: 10.1038/nrclinonc.2013.110 CrossRefPubMedGoogle Scholar
  2. 2.
    Diaz LA, Bardelli A (2014) Liquid biopsies: genotyping circulating tumor DNA. J Clin Oncol 32(6):579–586. doi: 10.1200/jco.2012.45.2011 CrossRefPubMedGoogle Scholar
  3. 3.
    Stroun M, Lyautey J, Lederrey C, Olson-Sand A, Anker P (2001) About the possible origin and mechanism of circulating DNA apoptosis and active DNA release. Clin Chim Acta 313(1–2):139–142CrossRefPubMedGoogle Scholar
  4. 4.
    van der Vaart M, Pretorius PJ (2008) Circulating DNA. Its origin and fluctuation. Ann NY Acad Sci 1137:18–26. doi: 10.1196/annals.1448.022 CrossRefPubMedGoogle Scholar
  5. 5.
    Diehl F, Schmidt K, Choti MA, Romans K, Goodman S, Li M, Thornton K, Agrawal N, Sokoll L, Szabo SA, Kinzler KW, Vogelstein B, Diaz LA Jr (2008) Circulating mutant DNA to assess tumor dynamics. Nat Med 14(9):985–990. doi: 10.1038/nm.1789 CrossRefPubMedCentralPubMedGoogle Scholar
  6. 6.
    Lo YM, Zhang J, Leung TN, Lau TK, Chang AM, Hjelm NM (1999) Rapid clearance of fetal DNA from maternal plasma. Am J Hum Genet 64(1):218–224. doi: 10.1086/302205 CrossRefPubMedCentralPubMedGoogle Scholar
  7. 7.
    Schwarzenbach H (2013) Circulating nucleic acids as biomarkers in breast cancer. Breast Cancer Res 15(5):211. doi: 10.1186/bcr3446 CrossRefPubMedCentralPubMedGoogle Scholar
  8. 8.
    Comprehensive molecular portraits of human breast tumours (2012). Nature 490 (7418):61–70. doi: 10.1038/nature11412
  9. 9.
    Stephens PJ, Tarpey PS, Davies H, Van Loo P, Greenman C, Wedge DC, Nik-Zainal S, Martin S, Varela I, Bignell GR, Yates LR, Papaemmanuil E, Beare D, Butler A, Cheverton A, Gamble J, Hinton J, Jia M, Jayakumar A, Jones D, Latimer C, Lau KW, McLaren S, McBride DJ, Menzies A, Mudie L, Raine K, Rad R, Chapman MS, Teague J, Easton D, Langerod A, Lee MT, Shen CY, Tee BT, Huimin BW, Broeks A, Vargas AC, Turashvili G, Martens J, Fatima A, Miron P, Chin SF, Thomas G, Boyault S, Mariani O, Lakhani SR, van de Vijver M, van’t Veer L, Foekens J, Desmedt C, Sotiriou C, Tutt A, Caldas C, Reis-Filho JS, Aparicio SA, Salomon AV, Borresen-Dale AL, Richardson AL, Campbell PJ, Futreal PA, Stratton MR (2012) The landscape of cancer genes and mutational processes in breast cancer. Nature 486(7403):400–404. doi: 10.1038/nature11017 PubMedCentralPubMedGoogle Scholar
  10. 10.
    Samuels Y, Diaz LA Jr, Schmidt-Kittler O, Cummins JM, Delong L, Cheong I, Rago C, Huso DL, Lengauer C, Kinzler KW, Vogelstein B, Velculescu VE (2005) Mutant PIK3CA promotes cell growth and invasion of human cancer cells. Cancer Cell 7(6):561–573. doi: 10.1016/j.ccr.2005.05.014 CrossRefPubMedGoogle Scholar
  11. 11.
    Kang S, Bader AG, Vogt PK (2005) Phosphatidylinositol 3-kinase mutations identified in human cancer are oncogenic. Proc Natl Acad Sci USA 102(3):802–807. doi: 10.1073/pnas.0408864102 CrossRefPubMedCentralPubMedGoogle Scholar
  12. 12.
    Dawson SJ, Tsui DW, Murtaza M, Biggs H, Rueda OM, Chin SF, Dunning MJ, Gale D, Forshew T, Mahler-Araujo B, Rajan S, Humphray S, Becq J, Halsall D, Wallis M, Bentley D, Caldas C, Rosenfeld N (2013) Analysis of circulating tumor DNA to monitor metastatic breast cancer. N Engl J Med 368(13):1199–1209. doi: 10.1056/NEJMoa1213261 CrossRefPubMedGoogle Scholar
  13. 13.
    Murtaza M, Dawson SJ, Tsui DW, Gale D, Forshew T, Piskorz AM, Parkinson C, Chin SF, Kingsbury Z, Wong AS, Marass F, Humphray S, Hadfield J, Bentley D, Chin TM, Brenton JD, Caldas C, Rosenfeld N (2013) Non-invasive analysis of acquired resistance to cancer therapy by sequencing of plasma DNA. Nature 497(7447):108–112. doi: 10.1038/nature12065 CrossRefPubMedGoogle Scholar
  14. 14.
    Beaver JA, Jelovac D, Balukrishna S, Cochran RL, Croessmann S, Zabransky DJ, Wong HY, Valda Toro P, Cidado J, Blair BG, Chu D, Burns T, Higgins MJ, Stearns V, Jacobs L, Habibi M, Lange J, Hurley PJ, Lauring J, VanDenBerg DA, Kessler J, Jeter S, Samuels ML, Maar D, Cope L, Cimino-Mathews A, Argani P, Wolff AC, Park BH (2014) Detection of cancer DNA in plasma of patients with early-stage breast cancer. Clin Cancer Res 20(10):2643–2650. doi: 10.1158/1078-0432.ccr-13-2933 CrossRefPubMedGoogle Scholar
  15. 15.
    Bettegowda C, Sausen M, Leary RJ, Kinde I, Wang Y, Agrawal N, Bartlett BR, Wang H, Luber B, Alani RM, Antonarakis ES, Azad NS, Bardelli A, Brem H, Cameron JL, Lee CC, Fecher LA, Gallia GL, Gibbs P, Le D, Giuntoli RL, Goggins M, Hogarty MD, Holdhoff M, Hong SM, Jiao Y, Juhl HH, Kim JJ, Siravegna G, Laheru DA, Lauricella C, Lim M, Lipson EJ, Marie SK, Netto GJ, Oliner KS, Olivi A, Olsson L, Riggins GJ, Sartore-Bianchi A, Schmidt K, Shih IM, Oba-Shinjo SM, Siena S, Theodorescu D, Tie J, Harkins TT, Veronese S, Wang TL, Weingart JD, Wolfgang CL, Wood LD, Xing D, Hruban RH, Wu J, Allen PJ, Schmidt CM, Choti MA, Velculescu VE, Kinzler KW, Vogelstein B, Papadopoulos N, Diaz LA Jr (2014) Detection of circulating tumor DNA in early- and late-stage human malignancies. Sci Transl Med 6(224):224ra224. doi: 10.1126/scitranslmed.3007094 CrossRefGoogle Scholar
  16. 16.
    Goldhirsch A, Wood WC, Gelber RD, Coates AS, Thurlimann B, Senn HJ (2003) Meeting highlights: updated international expert consensus on the primary therapy of early breast cancer. J Clin Oncol 21(17):3357–3365. doi: 10.1200/jco.2003.04.576 CrossRefPubMedGoogle Scholar
  17. 17.
    Goldhirsch A, Glick JH, Gelber RD, Coates AS, Thurlimann B, Senn HJ (2005) Meeting highlights: international expert consensus on the primary therapy of early breast cancer 2005. Ann Oncol 16(10):1569–1583. doi: 10.1093/annonc/mdi326 CrossRefPubMedGoogle Scholar
  18. 18.
    Goldhirsch A, Wood WC, Gelber RD, Coates AS, Thurlimann B, Senn HJ (2007) Progress and promise: highlights of the international expert consensus on the primary therapy of early breast cancer 2007. Ann Oncol 18(7):1133–1144. doi: 10.1093/annonc/mdm271 CrossRefPubMedGoogle Scholar
  19. 19.
    Goldhirsch A, Ingle JN, Gelber RD, Coates AS, Thurlimann B, Senn HJ (2009) Thresholds for therapies: highlights of the St Gallen International Expert Consensus on the primary therapy of early breast cancer 2009. Ann Oncol 20(8):1319–1329. doi: 10.1093/annonc/mdp322 CrossRefPubMedCentralPubMedGoogle Scholar
  20. 20.
    Bloom HJ, Richardson WW (1957) Histological grading and prognosis in breast cancer; a study of 1409 cases of which 359 have been followed for 15 years. Br J Cancer 11(3):359–377CrossRefPubMedCentralPubMedGoogle Scholar
  21. 21.
    Fujita N, Nakayama T, Yamamoto N, Kim SJ, Shimazu K, Shimomura A, Maruyama N, Morimoto K, Tamaki Y, Noguchi S (2012) Methylated DNA and total DNA in serum detected by one-step methylation-specific PCR is predictive of poor prognosis for breast cancer patients. Oncology 83(5):273–282. doi: 10.1159/000342083 CrossRefPubMedGoogle Scholar
  22. 22.
    Becker S, Becker-Pergola G, Banys M, Krawczyk N, Wallwiener D, Solomayer E, Schuetz C, Fehm T (2009) Evaluation of a RT-PCR based routine screening tool for the detection of disseminated epithelial cells in the bone marrow of breast cancer patients. Breast Cancer Res Treat 117(2):227–233. doi: 10.1007/s10549-008-0174-3 CrossRefPubMedGoogle Scholar
  23. 23.
    Turner NC, Garcia-Murillas I, Schiavon G, Hrebien S, Osin P, Nerurkar A, Kozarewa I, Garrido JA, Dowsett M, Smith IE (2014) Tracking tumor-specific mutations in circulating-free DNA to predict early relapse after treatment of primary breast cancer. J Clin Oncol 32:511Google Scholar
  24. 24.
    Miller TW, Rexer BN, Garrett JT, Arteaga CL (2011) Mutations in the phosphatidylinositol 3-kinase pathway: role in tumor progression and therapeutic implications in breast cancer. Breast Cancer Res 13(6):224. doi: 10.1186/bcr3039 CrossRefPubMedCentralPubMedGoogle Scholar
  25. 25.
    Berns K, Horlings HM, Hennessy BT, Madiredjo M, Hijmans EM, Beelen K, Linn SC, Gonzalez-Angulo AM, Stemke-Hale K, Hauptmann M, Beijersbergen RL, Mills GB, van de Vijver MJ, Bernards R (2007) A functional genetic approach identifies the PI3 K pathway as a major determinant of trastuzumab resistance in breast cancer. Cancer Cell 12(4):395–402. doi: 10.1016/j.ccr.2007.08.030 CrossRefPubMedGoogle Scholar
  26. 26.
    Andre F, Bachelot T, Commo F, Campone M, Arnedos M, Dieras V, Lacroix-Triki M, Lacroix L, Cohen P, Gentien D, Adelaide J, Dalenc F, Goncalves A, Levy C, Ferrero JM, Bonneterre J, Lefeuvre C, Jimenez M, Filleron T, Bonnefoi H (2014) Comparative genomic hybridisation array and DNA sequencing to direct treatment of metastatic breast cancer: a multicentre, prospective trial (SAFIR01/UNICANCER). Lancet Oncol 15(3):267–274. doi: 10.1016/s1470-2045(13)70611-9 CrossRefPubMedGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2015

Authors and Affiliations

  • Chiya Oshiro
    • 1
  • Naofumi Kagara
    • 1
    Email author
  • Yasuto Naoi
    • 1
  • Masafumi Shimoda
    • 1
  • Atsushi Shimomura
    • 1
  • Naomi Maruyama
    • 1
  • Kenzo Shimazu
    • 1
  • Seung Jin Kim
    • 1
  • Shinzaburo Noguchi
    • 1
  1. 1.Department of Breast and Endocrine SurgeryOsaka University Graduate School of MedicineSuita-shiJapan

Personalised recommendations