Monitoring Breast Cancer Response to Treatment Using Stokes Shift Spectroscopy of Blood Plasma

  • Krishnamoorthy Chithra
  • Prakasarao Aruna
  • Gnanatheepam Einstein
  • Srinivasan Vijayaraghavan
  • Singaravelu GanesanEmail author


With the emerging trend of personalized cancer treatment, there is a need to develop noninvasive/minimally invasive techniques for treatment monitoring. In this regard, in this work fluorescence analysis of blood plasma of breast cancer patients has been used for the evaluation of response to treatment. This approach delivers information not only about the change in biochemical constituents but also about the altered metabolic pathway. Spectral deconvolution method is employed to compute the fluorescence intensity, peak wavelength, and full-width half maxima for different endogenous fluorophores. The fluorescence measurements were made on blood plasma collected from 10 normal subjects, 10 pre-treated cancer patients, and 10 post-treated patients. Besides, variations in relative concentration of tryptophan, collagen, NADH, and FAD, peak shifts and broadening of peaks are observed for tryptophan, NADH, and FAD, in blood plasma of pre-treated cancer patients indicating both biochemical and microenvironmental changes at cellular level. Further, the spectral profile of blood plasma of post-treated patients found to be similar to blood plasma of normal subjects. Linear discriminant analysis showed that pre-treated and post-treated breast cancer is discriminated with a sensitivity and specificity of 100% and 100% respectively.


Stokes shift spectroscopy Blood plasma Breast cancer Spectral deconvolution method Treatment monitoring 



Authors gratefully acknowledge DAE-BRNS (No.:2009/34/38/BRNS) for funding this work.

Compliance with Ethical Standards

Conflict of Interest

The authors declare that there is no conflict of interest regarding the publication of this article.


  1. 1.
    Siegel RL, Miller KD, Fedewa SA, Ahnen DJ, Meester RG, Barzi A, Jemal A (2017) Colorectal cancer statistics, 2017. CA: CA Cancer J Clin 67(3):177–193Google Scholar
  2. 2.
    Harris L, Fritsche H, Mennel R, Norton L, Ravdin P, Taube S, Somerfield MR, Hayes DF, Bast RC Jr (2007) American Society of Clinical Oncology 2007 update of recommendations for the use of tumor markers in breast cancer. J Clin Oncol 25(33):5287–5312Google Scholar
  3. 3.
    Erdi YE (2012) Limits of tumor detectability in nuclear medicine and PET. Mol Imaging Radionucl Ther 21(1):23Google Scholar
  4. 4.
    McCormack DR, Walsh AJ, Sit W, Arteaga CL, Chen J, Cook RS, Skala MC (2014) In vivo hyperspectral imaging of microvessel response to trastuzumab treatment in breast cancer xenografts. Biomed Opt Express 5(7):2247–2261Google Scholar
  5. 5.
    Tromberg BJ, Zhang Z, Leproux A, O'Sullivan TD, Cerussi AE, Carpenter PM, Mehta RS, Roblyer D, Yang W, Paulsen KD, Pogue BW (2016) Predicting responses to neoadjuvant chemotherapy in breast cancer: ACRIN 6691 trial of diffuse optical spectroscopic imaging. Cancer Res 76(20):5933–5944Google Scholar
  6. 6.
    Vidyasagar MS, Maheedhar K, Vadhiraja BM, Fernendes DJ, Kartha VB, Krishna CM (2008) Prediction of radiotherapy response in cervix cancer by Raman spectroscopy: a pilot study. Biopolymers 89(6):530–537Google Scholar
  7. 7.
    Tsuyuki S, Yamaguchi A, Kawata Y, Kawaguchi K (2015) Assessing the effects of neoadjuvant chemotherapy on lymphatic pathways to sentinel lymph nodes in cases of breast cancer: usefulness of the indocyanine green-fluorescence method. Breast 24(3):298–301Google Scholar
  8. 8.
    Pu Y, Tang GC, Wang WB, Savage HE, Schantz SP, Alfano RR (2011) Native fluorescence spectroscopic evaluation of chemotherapeutic effects on malignant cells using nonnegative matrix factorization analysis. Technol Cancer Res Treat 10(2):113–120Google Scholar
  9. 9.
    Pu Y, Tang G, Wang W, Savage HE, Schantz SP, Alfano RR (2012) Chemotherapeutic effects on breast malignant cells evaluated by native fluorescence spectroscopy. In: Biomedical optics JM3A-45Google Scholar
  10. 10.
    Chithra K, Vijayaraghavan S, Prakasarao A, Singaravelu G (2017) Study of anti-cancer effects of chemotherapeutic agents and radiotherapy in breast cancer patients using fluorescence spectroscopy. In: Optical biopsy XV: toward real-time spectroscopic imaging and diagnosis, vol 10060, p 100600LGoogle Scholar
  11. 11.
    Sivabalan S, Vedeswari CP, Jayachandran S, Koteeswaran D, Pravda C, Aruna P, Ganesan S (2010) In vivo native fluorescence spectroscopy and nicotinamide adinine dinucleotide/flavin adenine dinucleotide reduction and oxidation states of oral submucous fibrosis for chemopreventive drug monitoring. J Biomed Opt 15(1):017010Google Scholar
  12. 12.
    Han SH, Song TK (2015) In vivo fluorescence spectroscopic monitoring of radiotherapy in cancer treatment. Int J Cancer Ther Oncol 3(1):03013Google Scholar
  13. 13.
    Brown JQ, Bydlon TM, Richards LM, Yu B, Kennedy SA, Geradts J, Wilke LG, Junker MK, Gallagher J, Barry WT, Ramanujam N (2010) Optical assesssment of tumor resection margins in the breast. IEEE J Sel Top Quantum Electron 16(3):530–544Google Scholar
  14. 14.
    Alfano RR, Tata DB, Cordero J, Tomashefsky P, Longo F, Alfano M (1984) Laser induced fluorescence spectroscopy from native cancerous and normal tissue. IEEE J Quantum Electron 20(12):1507–1511Google Scholar
  15. 15.
    Zhadin NN, Yang Y, Ganesan S, Ockman N, Alfano RR (1996) Enhancement of the fluorescence cancer diagnostic method of tissues using diffuse reflectance and the analysis of oxygenation state. In: Advances in laser and light spectroscopy to diagnose cancer and other diseases III: optical biopsy, vol 2679, pp 142–149Google Scholar
  16. 16.
    Ganesan S, Sacks PG, Yang Y, Katz A, Al-Rawi M, Savage HE, Schantz SP, Alfano RR (1998) Native fluorescence spectroscopy of normal and malignant epithelial cells. Cancer Biochem Biophys 16(4):365–373Google Scholar
  17. 17.
    Liu Q, Grant G, Vo-Dinh T (2010) Investigation of synchronous fluorescence method in multicomponent analysis in tissue. IEEE J Sel Top Quantum Electron 16(4):927–940Google Scholar
  18. 18.
    Majumder SK (2000) Synchronous luminescence spectroscopy for oral cancer diagnosis. Lasers Life Sci 9:143–152Google Scholar
  19. 19.
    Vengadesan N, Anbupalam T, Hemamalini S, Ebenezar J, Muthvelu K, Koteeswaran D, Aruna PR, Ganesan S (2002) Characterization of cervical normal and abnormal tissues by synchronous luminescence spectroscopy. In Optical Biopsy IV 4613:13–18Google Scholar
  20. 20.
    Alfano RR, Yang Y (2003) Stokes shift emission spectroscopy of human tissue and key biomolecules. IEEE J Sel Top Quantum Electron 9(2):148–153Google Scholar
  21. 21.
    Dramicanin T, Dramicanin MD, Dimitrijevic B, Jokanovic V, Lukic S (2006) Discrimination between normal and malignant breast tissues by synchronous luminescence spectroscopy. Acta Chim Slov 53(4):444Google Scholar
  22. 22.
    Ebenezar J, Aruna P, Ganesan S (2010) Synchronous fluorescence spectroscopy for the detection and characterization of cervical cancers in vitro. Photochem Photobiol 86(1):77–86Google Scholar
  23. 23.
    Pu Y, Wang W, Yang Y, Alfano RR (2013) Stokes shift spectroscopic analysis of multifluorophores for human cancer detection in breast and prostate tissues. J Biomed Opt 18(1):017005Google Scholar
  24. 24.
    Muthuvelu K, Shanmugam S, Koteeswaran D, Srinivasan S, Venkatesan P, Aruna P, Ganesan S (2011) Synchronous luminescence spectroscopic characterization of blood elements of normal and patients with cervical cancer. In: Optical biopsy IX, vol 7895, p 78950F International Society for Optics and PhotonicsGoogle Scholar
  25. 25.
    Rajasekaran R, Aruna P, Koteeswaran D, Baludavid M, Ganesan S (2014) Synchronous luminescence spectroscopic characterization of urine of normal subjects and cancer patients. J Fluoresc 24(4):1199–1205Google Scholar
  26. 26.
    Kumar P, Ashutosh K, Pradhan A (2018) Comparative study between diagnostic mediums: human tissue and saliva for oral cancer detection using stokes shift spectroscopy. In: Optics in Health Care and Biomedical Optics VIII, vol 10820, p 108200L International Society for Optics and PhotonicsGoogle Scholar
  27. 27.
    Yuvaraj M, Aruna P, Koteeswaran D, Muthuvelu K, Ganesan S (2018) UV-native fluorescence steady and excited state kinetics of salivary protein of normal subjects, oral premalignant and malignant conditions. J Lumin 196:236–243Google Scholar
  28. 28.
    Xu X, Meng J, Hou S, Ma H, Wang D (1988) The characteristic fluorescence of the serum of cancer patients. J Lumin 40:219–220Google Scholar
  29. 29.
    Hubmann MR, Leiner MJ, Schaur RJ (1990) Ultraviolet fluorescence of human sera: I. sources of characteristic differences in the ultraviolet fluorescence spectra of sera from normal and cancer-bearing humans. Clin Chem 36(11):1880–1883Google Scholar
  30. 30.
    Madhuri S, Vengadesan N, Aruna P, Koteeswaran D, Venkatesan P, Ganesan S (2003) Native fluorescence spectroscopy of blood plasma in the characterization of Oral malignancy. Photochem Photobiol 78(2):197–204Google Scholar
  31. 31.
    Chauhan P, Yadav R, Kaushal V, Beniwal P (2016) Evaluation of serum biochemical profile of breast cancer patients. Int J Med Res Health Sci 5(7):1Google Scholar
  32. 32.
    Gnanatheepam E, Kanniyappan U, Dornadula K, Prakasarao A, Singaravelu G (2019) Synchronous luminescence spectroscopy as a tool in the discrimination and characterization of Oral Cancer tissue. J Fluoresc:1–7Google Scholar
  33. 33.
    Yang Y, Katz A, Celmer EJ, Zurawska-Szczepaniak M, Alfano RR (1997) Fundamental differences of excitation spectrum between malignant and benign breast tissues. Photochem Photobiol 66(4):518–522Google Scholar
  34. 34.
    Cascino A, Cangiano C, Ceci F, Franchi F, Mineo T, Mulieri M, Muscaritoli M, Rossi FF (1991) Increased plasma free tryptophan levels in human cancer: a tumor related effect? Anticancer Res 11(3):1313–1316Google Scholar
  35. 35.
    Miyagi Y, Higashiyama M, Gochi A, Akaike M, Ishikawa T, Miura T, Saruki N, Bando E, Kimura H, Imamura F, Moriyama M (2011) Plasma free amino acid profiling of five types of cancer patients and its application for early detection. PLoS One 6(9):e24143Google Scholar
  36. 36.
    Zhang L, Pu Y, Xue J, Pratavieira S, Xu B, Achilefu S, Alfano RR (2014) Tryptophan as the fingerprint for distinguishing aggressiveness among breast cancer cell lines using native fluorescence spectroscopy. J Biomed Opt 19(3):037005Google Scholar
  37. 37.
    Brisson BK, Mauldin EA, Lei W, Vogel LK, Power AM, Lo A, Dopkin D, Khanna C, Wells RG, Puré E, Volk SW (2015) Type III collagen directs stromal organization and limits metastasis in a murine model of breast cancer. Am J Pathol 185(5):1471–1486Google Scholar
  38. 38.
    Schomacker KT, Frisoli JK, Compton CC, Flotte TJ, Richter JM, Nishioka NS, Deutsch TF (1992) Ultraviolet laser-induced fluorescence of colonic tissue: basic biology and diagnostic potential. Lasers Surg Med 12(1):63–78Google Scholar
  39. 39.
    Pandey K, Pradhan A, Agarwal A, Bhagoliwal A, Agarwal N (2012) Fluorescence spectroscopy: a new approach in cervical cancer. J Obstet Gynaecol India 62(4):432–436Google Scholar
  40. 40.
    Mazouni C, Arun B, Andre F, Ayers M, Krishnamurthy S, Wang B, Hortobagyi GN, Buzdar AU, Pusztai L (2008) Collagen IV levels are elevated in the serum of patients with primary breast cancer compared to healthy volunteers. Br J Cancer 99(1):68Google Scholar
  41. 41.
    Miller JA, Pappan K, Thompson PA, Want EJ, Siskos AP, Keun HC, Wulff J, Hu C, Lang JE, Chow HHS (2015) Plasma metabolomic profiles of breast cancer patients after short-term limonene intervention. Cancer Prev Res 8(1):86–93Google Scholar
  42. 42.
    Chance B, Schoener B, Oshino R, Itshak F, Nakase Y (1979) Oxidation-reduction ratio studies of mitochondria in freeze-trapped samples. NADH and flavoprotein fluorescence signals. J Biol Chem 254(11):4764–4771Google Scholar
  43. 43.
    Pradhan A, Pal P, Durocher G, Villeneuve L, Balassy A, Babai F, Gaboury L, Blanchard L (1995) Steady state and time-resolved fluorescence properties of metastatic and non-metastatic malignant cells from different species. J Photochem Photobiol B 31(3):101–112Google Scholar
  44. 44.
    Glassman WS, Liu CH, Tang GC, Lubicz S, Alfano R (1992) Ultraviolet excited fluorescence spectra from non-malignant and malignant tissues of the gynecological tract. Lasers Life Sci 5(1–2):49–58Google Scholar
  45. 45.
    Ostrander JH, McMahon CM, Lem S, Millon SR, Brown JQ, Seewaldt VL, Ramanujam N (2010) Optical redox ratio differentiates breast cancer cell lines based on estrogen receptor status. Cancer Res 70(11):4759–4766Google Scholar
  46. 46.
    Wojcieszyńska D, Hupert-Kocurek K, Guzik U (2012) Flavin-dependent enzymes in cancer prevention. Int J Mol Sci 13(12):16751–16768Google Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  • Krishnamoorthy Chithra
    • 1
    • 2
  • Prakasarao Aruna
    • 1
  • Gnanatheepam Einstein
    • 1
  • Srinivasan Vijayaraghavan
    • 2
  • Singaravelu Ganesan
    • 1
    Email author
  1. 1.Department of Medical PhysicsAnna UniversityChennaiIndia
  2. 2.Department of OncologyPaterson Cancer CenterChennaiIndia

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